U.S. patent application number 11/151147 was filed with the patent office on 2006-01-05 for screening arrangement for screening immunoassay tests and agglutination tests.
Invention is credited to Dene Baldwin, Christopher William Hand, Osborn Pierce Jones, Robin James Spivey.
Application Number | 20060003396 11/151147 |
Document ID | / |
Family ID | 10835523 |
Filed Date | 2006-01-05 |
United States Patent
Application |
20060003396 |
Kind Code |
A1 |
Spivey; Robin James ; et
al. |
January 5, 2006 |
Screening arrangement for screening immunoassay tests and
agglutination tests
Abstract
Immunoassay tests or agglutination tests run on samples of
bodily fluid to detect the presence of particular compounds such as
drugs in the body may be screened in the screening device. A test
membrane is inserted into the screening device and illuminated. The
reflected image is detected and the digitised data processed. For
immunoassay tests, the digitised data is segmented and data for the
test region is compared to that from the control region and the
background regions to determine whether the test data exhibits any
significant results. For agglutination tests, the digitised data is
processed to determine the number and size of the areas of
coagulation to determine whether the test data exhibits any
significant results. A swab for taking a bodily sample incorporates
a run fluid capsule. Once an adequate sample of bodily fluid has
been collected and the swab is placed in contact with the test
membrane, the run fluid capsule is pierced by a spike provided on
the swab, the run fluid mixes with the sample and the mixture is
conveyed to the test membrane. Alternatively or in addition the
swab has a main tube and a capillary tube. A run detector in the
capillary tube detects when an adequate sample of bodily fluid has
been taken.
Inventors: |
Spivey; Robin James;
(Bangor, GB) ; Hand; Christopher William;
(Abingdon, GB) ; Baldwin; Dene; (Oxford, GB)
; Jones; Osborn Pierce; (Gwernafalau, GB) |
Correspondence
Address: |
FLYNN THIEL BOUTELL & TANIS, P.C.
2026 RAMBLING ROAD
KALAMAZOO
MI
49008-1631
US
|
Family ID: |
10835523 |
Appl. No.: |
11/151147 |
Filed: |
June 13, 2005 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09760374 |
Jan 12, 2001 |
|
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|
11151147 |
Jun 13, 2005 |
|
|
|
PCT/GB99/02261 |
Jul 14, 1999 |
|
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09760374 |
Jan 12, 2001 |
|
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Current U.S.
Class: |
435/7.92 ;
435/287.2 |
Current CPC
Class: |
Y10S 435/97 20130101;
G01N 33/54366 20130101; Y10S 435/805 20130101; Y10S 435/81
20130101; Y10S 436/805 20130101 |
Class at
Publication: |
435/007.92 ;
435/287.2 |
International
Class: |
C12Q 1/68 20060101
C12Q001/68; G01N 33/53 20060101 G01N033/53; G01N 33/537 20060101
G01N033/537; G01N 33/543 20060101 G01N033/543; C12M 1/34 20060101
C12M001/34 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 14, 1998 |
GB |
9815302.6 |
Claims
1-17. (canceled)
18. A screening device for interpreting the results of an
agglutination test, said test having areas of coagulation and
background areas, said device comprising: a light source for
illuminating said test; an array of photosensitive detectors for
detecting the intensity of light from said light source which is
reflected from said areas of coagulation and background areas of
said agglutination test; a digitiser, coupled to the output of said
array of photosensitive detectors, for representing said intensity
of the detected light by a data array; a threshold processor,
coupled to the output of said digitiser, for thresholding said
digitised data to distinguish between background areas and areas of
coagulation; a first data processor, coupled to said threshold
processor, for identifying from said thresholded data areas of
coagulation and estimating the size of said areas of coagulation; a
second data processor, coupled to the output of said first data
processor, for determining whether said areas of coagulation
exhibit a statistically significant result; and an output, coupled
to said second data processor, for outputting the results from said
second data processor.
19. A screening device according to claim 18, wherein a noise
reduction processor is coupled between the output of said digitiser
and said first data processor for performing noise reduction on
said digitised data.
20. A method of screening an agglutination test, said test having
areas of coagulation and background areas, said method comprising
the steps of: illuminating said test; detecting the intensity of
light which is reflected from said areas of coagulation and
background areas of said agglutination test; representing the
intensity of said detected light in a data array; thresholding said
data array to distinguish between background areas and areas of
coagulation; identifying areas of coagulation and estimating the
number of said areas of coagulation; determining whether said areas
of coagulation exhibit a statistically significant result; and
outputting said results.
21. A method according to claim 20, wherein noise reduction of said
digitised data is performed prior to thresholding said digitised
data.
22. A swab for taking a sample of bodily fluid and transferring
said sample to a test strip, said swab comprising: a collection
pad, a tube, a run fluid chamber and an elongate spike; said tube
having a bore running axially along its length and having open
upper and lower ends, said lower end of said tube being disposed
axially adjacent to and in communication with said collection pad;
said run fluid chamber being located in said bore of said tube; and
said elongate spike being moveable in said bore of said tube from a
sampling position to a sample transferring position, said elongate
spike being axially remote from said run fluid chamber in said
sampling position and piercingly contacting said run fluid chamber
in said sample transferring position.
23. A swab according to claim 22, wherein a detector is provided
for detecting that an adequate sample of bodily fluid has been
collected.
24. A swab according to claim 23, wherein said detector is located
in said tube and comprises a dye release pad and a dye receptor
pad, said dye release pad being positioned in closer axial
proximity to said collection pad than said dye receptor pad, and
said tube being substantially transparent in the vicinity of said
dye receptor pad.
25. A swab according to claim 24, wherein a filter pad is located
axially between said collection pad and said dye release pad.
26. A swab according to claim 22, wherein an outer tube is
provided, said outer tube being open at the lower end thereof and
housing said upper end of said tube; said elongate spike being held
captive between said tube and said outer tube, said outer tube
being moveable relative to said tube whereby movement of said outer
tube relative to said tube causes said elongate spike to move from
said sampling position to said sample transferring position.
27. A swab for taking a sample of bodily fluid and transferring
said sample to a test strip, said swab comprising: a collection
pad, a main tube, a run detector and a capillary tube; said main
tube having an axial bore and being open at the lower end thereof,
said lower end of said tube being disposed axially adjacent to and
in communication with said collection pad; and said capillary tube
being open at the upper and lower ends thereof, said open lower end
of said capillary tube being in communication with said collection
pad and said run detector being located in said capillary tube, the
distance between said run detector and said collection pad being
such that said run detector detects when an adequate sample of
bodily fluid has been collected.
28. A swab according to claim 27, wherein the wall of said main
tube defines a port, said open lower end of said capillary tube
being connected to said port and communicating with said main
tube.
29. A swab according to claim 28, wherein said run detector
comprises a dye release pad and a dye receptor pad, said dye
release pad being positioned in closer proximity to said lower end
of said capillary tube than said dye receptor pad.
30. A swab according to claim 27, wherein a pierceable run fluid
chamber is located in said main tube.
Description
RELATIONSHIP TO EARLIER FILED APPLICATION
[0001] This application is a continuation-in-part of International
Application PCT/GB99/02261, publication No. WO 00/04381, with an
International filing date of Jul. 14, 1999.
BACKGROUND OF THE INVENTION
[0002] This invention relates to a screening device and methods of
screening immunoassay tests and agglutination tests. In particular
the invention is applicable to a screening device for detecting the
presence and concentration of particular drugs in a sample of
saliva.
[0003] Samples of bodily fluid such as blood, sweat, urine and
saliva may be used to detect the presence of particular compounds,
such as drugs, in the body. Known methods of testing such samples
for the presence of compounds include immunoassay "strip" testing
where an antibody is labelled with a suitable marker, for example a
visible marker such as colloidal gold, and drawn along a membrane
passing over test regions and a control region impregnated with
analyte conjugate substances or other binding substances. The
presence of particular compounds in the sample are detected by a
visible change occurring in the corresponding region due to the
interaction of the labelled antibodies and the conjugate substances
resulting in visible lines forming on the membrane in some of these
regions. The colour formed may be proportional to or inversely
proportional to analyte concentration depending on the assay
format.
[0004] The interpretation of the lines formed by such immunoassay
testing has previously been carried out subjectively by an operator
comparing the intensity of the test line (or the absence or
presence of a line) with that of a control, or reference, line.
[0005] U.S. Pat. No. 5,580,794 describes a disposable electronic
assay device. For single analytes only one light source and
detector is necessary; for two analytes, two sets of light source
and detector is necessary and so on.
SUMMARY OF THE INVENTION
[0006] The invention in its various aspects is defined in the
independent claims below, to which reference should now be made,
Advantageous features are set forth in the appendant claims.
[0007] We have appreciated that in some fields of drug testing, for
example in the use of a sample matrix other than urine such as
saliva or blood, the amount of drug present in the sample may be
very low, and the operator must be able to distinguish between a
negative test corresponding to a complete absence of the drug in
the sample and very low levels of drug present. This is difficult,
requiring highly trained and skilled operators, and can prove
unreliable when the levels of drugs are very low. For example, it
is particularly difficult for an operator to distinguish between
levels of cannabis of 6 ng/mL or lower by eye. If the test is to be
run outside the laboratory, it is even more likely to be subject to
inaccuracies which may be exacerbated by poor lighting conditions
or by other environmental factors. We have, therefore, recognised
the need for a portable drug tester which produces reliable and
reproducible results.
[0008] We have also appreciated that a non-invasive test that can
be conducted for example by the roadside would be beneficial. In a
preferred embodiment of the invention we have therefore provided an
automatic drug tester which can detect even very low levels of
drugs from a saliva sample.
[0009] Preferred embodiments of the invention are described with
respect to the drawings. In two of the preferred embodiments,
immunoassay tests and agglutination tests run on samples of bodily
fluid to detect the presence of particular compounds such as drugs
in the body may be screened in the screening device. A test
membrane is inserted into the screening device and illuminated. The
reflected image is detected and the digitised data processed. For
immunoassay tests, the digitised data is segmented and data for the
test region is compared to that from the control region and the
background regions to determine whether the test data exhibits any
significant results. For agglutination tests, the digitised data is
processed to determine the number and size of the areas of
coagulation to determine whether the test data exhibits any
significant results.
[0010] In another preferred embodiment, a swab for taking a bodily
sample incorporates a run fluid capsule. Once an adequate sample of
bodily fluid has been collected and the swab is placed in contact
with the test membrane, the run fluid capsule is pierced by a spike
provided on the swab, the run fluid mixes with the sample and the
mixture is conveyed to the test membrane. In another preferred
embodiment, the swab has a main tube and a capillary tube. A run
detector in the capillary tube detects when an adequate sample of
bodily fluid has been taken.
BRIEF DESCRIPTION OF THE DRAWINGS
[0011] Preferred embodiments of the invention will now be described
in more detail, by way of example, with reference to the drawings
in which:
[0012] FIG. 1 is an isometric view of a test cartridge;
[0013] FIG. 2 is a side view of an immunoassay test strip;
[0014] FIG. 3 is an isometric view of a preferred screening device
embodying the invention with a test swab and test cartridge located
ready for analysis;
[0015] FIG. 4 is a side view of the screening device, test
cartridge and test swab of FIG. 3;
[0016] FIG. 5 is a block diagram of the electrical controls and
electrical apparatus used in the screening device;
[0017] FIG. 6 shows a graph of the typical variation of pixel
intensity with pixel position for a single test and a single
reference test;
[0018] FIG. 7 is a block diagram of the electrical controls and
electrical apparatus which may alternatively or additionally be
used in the screening device;
[0019] FIG. 8 is a schematic view of the detected intensity of a
test strip showing the typical appearance of a control and test
zone after a test has been run;
[0020] FIG. 9 is a cut away side view of a second embodiment of a
test cartridge;
[0021] FIG. 10 is a cut away side view of a second embodiment of a
test swab;
[0022] FIG. 11 shows the cut away test swab of FIG. 10 inserted
into a test cartridge and deployed for running a test;
[0023] FIG. 12 is a cut away side view of a third embodiment of a
test swab;
[0024] FIG. 13 is a plan view of the test swab of FIG. 12;
[0025] FIG. 14 is a side view of the piercing spike for attachment
to a test cartridge for use with the test swab of FIGS. 12 and
13;
[0026] FIG. 15 is a plan view of the spike of FIG. 14;
[0027] FIG. 16 is a flow chart showing the operation of a second
embodiment of the screening device;
[0028] FIG. 17 is a schematic diagram showing the correspondence
between the sequence of the element-wise processing of the
monochrome data array and the position in the acquired image;
[0029] FIG. 18 is a flow chart showing the primary scan operation
of FIG. 16;
[0030] FIG. 19 is a flow chart showing the secondary scan operation
of FIG. 16;
[0031] FIG. 20 is a flow chart showing the tag tree compaction and
simplification operation of FIG. 16;
[0032] FIG. 21 is a plan view of a single reaction agglutination
test cartridge; and
[0033] FIG. 22 is a plan view of a multiple reaction agglutination
test cartridge.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
[0034] FIG. 1 shows a test cartridge 10 used to run the immunoassay
tests to be screened by the screening device. The test cartridge 10
may be disposable and is formed from a top 12 and a base 14. The
top 12 of the test cartridge 10 has a cylindrical swab holder 16
extending vertically from one of the shorter ends of an elongate
tray 18. The swab holder 16 is open at both ends.
[0035] Ridges 20 extend upwardly from both of the longer sides of
the elongate tray along the length of the tray. A rectangular
window 22 extends transversely between the ridges 20 across the
elongate tray and extends over a longitudinal length of the
elongate tray which is less than the overall length of the elongate
tray such that the window 22 is bounded on all four sides by the
elongate tray 18. The window 22 extends through the entire
thickness of the elongate tray 18.
[0036] FIG. 2 shows an immunoassay test strip 23. The upper surface
of a flat, elongate nitrocellulose membrane 24 is bonded to a waste
pad 28 at one end and to a conjugate release pad 27 at its other
end. Both the conjugate release pad 27 and the waste pad 28 overlap
the ends of the nitrocellulose membrane 24. The other end of the
conjugate release pad 27 overlaps an absorbent sample pad 26 and is
bonded at its upper surface to the lower surface of the absorbent
sample pad 26. When fluid is applied to the sample pad 26 it is
drawn along the sample pad by capillary action, through the
conjugate release pad 27 and nitrocellulose membrane 24 and surplus
fluid is absorbed by the waste pad 28.
[0037] The base 14 of the test cartridge 10 has a rectangular
portion with a rounded portion at one end. An immunoassay test
strip 23 is laid onto the upper surface of the base 14 with the
sample pad 26 located in the rounded portion of the base 14. The
immunoassay strip 23 (shown in dashed lines on FIG. 1) extends
longitudinally along the length of the base 14 from the end of the
base furthest from the rounded portion stopping within the rounded
portion but short of the end of the rounded portion. The top 12 is
then assembled onto the base 14 by fitting the cylindrical swab
holder 16 onto the rounded portion of the base 14 and the elongate
tray 18 of the top 12 onto the rectangular portion of the base 14.
The top 12 and base 14 are joined for example by gluing.
Alternatively, the top 12 may be designed to snap-fit onto the base
14. The top 12 may be made of a single unit so that the elongate
tray 18 and the swab holder 16 are a single piece.
[0038] The conjugate release pad 21 holds a mobile and visible
label, or marker, such as colloidal gold, and is in contact with
the nitro-cellulose membrane 24 such that when fluid is added to
the swab holder 16, it is drawn by capillary action downstream from
the swab holder 16 through the absorbent sample pad 26 through the
conjugate release pad 27 and subsequently through the
nitro-cellulose membrane 24. The use of cartridges of this type is
known in the prior art for example from EP 0291 194 by Unilever NV
titled "Immunoassays and devices therefor".
[0039] At discrete intervals along the nitro-cellulose membrane 24
drug-protein derivatives are biochemically bound to the
nitro-cellulose membrane, producing an immobile zone of
drug-protein derivative which spans the width of the
nitro-cellulose membrane. Towards the extreme downstream end of the
nitro-cellulose member, downstream of all the immobile drug-protein
derivative zones, is a control zone which also spans the width of
the nitro-cellulose membrane. The test zones and control zone are
interposed between background zones where the nitrocellulose
membrane 24 does not have bound drug conjugate but has been blocked
by other protein or other substances to prevent non-specific
binding. Antibodies to each drug which is to be tested for,
conjugated with colloidal gold, are placed on the conjugate release
pad 27. When saliva is transferred from the swab in the presence of
a run-fluid, the resulting sample passes across the absorbent
sample pad 26 and across the conjugate release pad 27 where it
mixes with the antibody-gold conjugates. The sample then travels
the length of the nitro-cellulose membrane 24.
[0040] If the particular drug is present in the sample it will bind
to the antibody-gold conjugate. When the bound drug subsequently
passes over the specific drug-protein derivative the antibody-gold
conjugate has already been bound to the drug in the sample and is
not free to bind with the drug-protein derivative bonded to the
membrane. If the particular drug is absent from the sample, the
antibody-gold conjugate will be free to bind to the drug-protein
conjugate causing the antibody-gold conjugate to become immobilised
at the site of the drug-protein conjugate. The visible marker is
deposited in the test zone as a coloured line or stripe. In between
these two extremes some of the antibody-gold conjugate will bind
with the drug-protein derivatives on the strip creating an
intermediate intensity of colour. The intensity of the colour on
the particular drug-protein zone is therefore inversely
proportional to the amount of drug present in the sample.
[0041] The depth of colour of the control zone should always be
significant and the control zone is designed with this in mind. The
colour of the control zone can then be used to indicate that the
test has been successfully run and to threshold colour levels in
specific drug conjugate zones.
[0042] FIG. 3 shows the test cartridge 10 of FIG. 1 located into
the screening device 30. The screening device 30 includes a
receiving section, an imaging section and a display section. The
receiving section is located at the rear of the screening device
and receives and aligns a test cartridge prior to the screening
operation. The imaging section is located centrally in the
screening device between the receiving section and the display
section and includes the illuminating and imaging equipment, the
processing capabilities and battery pack. At the front of the
screening device is the display section for outputting the results
of the screening operation. A cover (not shown) which is open at
the front end of the screening device 30 encases the remaining five
sides of the screening device 30. A facia cover (not shown) is
attached to the cover to completely encase the screening device 30,
protecting the screening device and the user from accidental
damage.
[0043] The receiving section includes a receiving bracket 32 and a
microswitch 43 and also positions and supports a half silvered
mirror 40 which forms part of the imaging section. The receiving
bracket 32 has a back 38 and two parallel arms 36. The back 38 is
connected at either end to one end of each arm forming a U-shaped
bracket. The open end of the U-shaped receiving bracket 32 is
directed outwardly from the screening device 30 and is aligned with
an opening in one side of the cover (not shown) towards the rear of
the screening device 30. The opening is large enough to allow a
test cartridge 10 to be inserted into the screening device 30. The
arms of the receiving bracket 32 are spaced apart by a distance
equal to the width of the test cartridge 10 and have the same
longitudinal length as that of the elongate tray 18 of the test
cartridge 10. The arms 36 have a C-shaped cross-section. When a
test cartridge is inserted into the opening the ridges 20 on the
test cartridge 10 mesh with the C-shaped cross-section of the arms
36 of the receiving bracket 32 to direct the test cartridge into
the screening device 30. The test cartridge 10 is inserted into the
screening device 30 until the end of the test cartridge reaches the
back of the receiving bracket when pressure against further
insertion will be felt.
[0044] A half silvered mirror 40, which forms part of the imaging
section of the screening device, is supported above the window 22
of the test cartridge 10 by a column 42 extending upwardly from the
arm 36 of the receiving bracket 32 nearest the rear of the
screening device 30.
[0045] A test swab 70, holding a saliva sample, is located in the
swab holder 16 of a disposable test cartridge 10. The test
cartridge 10 is inserted into the screening device 30 by the end
furthest from the swab holder 16 and is positively located in the
correct screening position by receiving bracket 32. The back 38 of
the receiving bracket 32 prevents the test cartridge 10 from being
inserted too far into the screening device 30 and ensures that the
window 22 of the cartridge 10 is located directly in front of and
beneath the CCD 34 of the screening device 30. Electrical circuitry
50 controlling the operation of the screening device including
operation of the CCD 34 are housed within the screening device 30
towards the front of the screening device 30 in front of the CCD
34, test cartridge 10, and rechargeable batteries 48.
[0046] A microswitch 43 is supported above the test cartridge 10
from the arm 36 of the receiving bracket 32 nearest the CCD 34.
When a test cartridge 10 is inserted into the screening device 30
the microswitch 43 is displaced vertically causing an electrical
signal to be emitted from the microswitch to signal that the
correct insertion of a test cartridge 10 has been detected. During
screening of the test cartridge 10, the microswitch 43 may resist
any displacement of the test cartridge 10 once it has been fully
inserted into the screening device.
[0047] The imaging section includes illuminating means,
photosensitive detector means, means for representing the intensity
of the detected light by a data array, data processing means for
segmenting the data and comparing the segmented data and output
means. The illuminating means is provided by three light emitting
devices (LEDs) 44 which are mounted in a horizontal line parallel
to the longitudinal length of the test cartridge 10 with the middle
LED centred vertically above the centre of window 22 of the test
cartridge 10. The photosensitive detector means and means for
representing the intensity of the detected light by a data array
are provided by a CCD 34 which includes an imager 82, a video
digitiser 84 and a video data interface 86 (shown on FIG. 5).
Alternatively, the photosensitive detector means may be made up
from a CCD array device together with a control and data conversion
interface. The imager of the CCD 34 is directed towards the rear of
the screening device 30. A mounting plate 46 is attached to the
upper body of the CCD 34 towards the front of the screening device
30. The mounting plate 46 extends horizontally from the body of the
CCD 34 towards the rear of the screening device 30 and finishes
directly above the window 22 of the test cartridge 10. Three LEDs
44 are attached in a row at the front of the underside of the
mounting plate 46. When illuminated, the light from the LEDs 44
shines directly onto the window 22 of the test cartridge 10. The
mirror 40 is inclined from the vertical by approximately 35.degree.
such that the window 22 of the test cartridge is reflected into the
field of view of the CCD 34. Light reflected from the immunoassay
test is detected by an array of photosensitive detectors in the
imager 82. The photosensitive detectors emit an electrical signal
proportional to the intensity of light detected. The video
digitiser 84 scans each of the photosensitive detectors in turn,
converting the analogue data to digital data and storing the data
in an array. The array of digital data is subsequently outputted to
a central processor unit (CPU) 80 via the video data interface
86.
[0048] Rechargeable batteries 48 supply power to the CCD 34, LEDs
44, microswitch 43 and electrical circuitry 50. The rechargeable
batteries 48 towards the front of the imaging section below the CCD
34. The electrical circuitry 50 which forms the final part of the
imaging section is described later with reference to FIG. 4 and
FIG. 7.
[0049] At the front of the screening device 30 is the display
section including two test indicator LEDs 52 and 54, a liquid
crystal device (LCD) 56, operating buttons 58 and 60 and a front
plate 62. The front plate 62 is slightly s.5 smaller than the facia
cover and is located at the front of the screening device 30
directly behind the facia cover. The two test indicator LEDs 52 and
54 are mounted at the top of the rear of the front plate with the
LEDs 52 and 54 protruding above the level of the front plate 62.
Holes in the top of the cover at its front corner allow the test
indicator LEDs 52 and 54 to protrude through the cover such that
they are visible on top of the device.
[0050] An LCD 56 and its associated backlight driver 94 are mounted
at the top of the front plate 62 between the front plate 62 and the
facia cover. The facia cover has a window through which the LCD 56
is visible but which obscures the backlight driver 94, located
behind the LCD 56, from view. Also mounted onto the front plate 62
between the facia cover and the front plate are two operating
buttons 58 and 60. The facia cover has holes in corresponding
locations to allow the user to operate the buttons 58 and 60
through the facia cover.
[0051] Additionally holes for an infra-red communication port and a
serial and parallel link for connecting the screening device to a
personal computer (PC) may be provided in the cover and
corresponding connections from the electrical circuitry 50 may be
provided.
[0052] FIG. 4 shows a partially sectioned side view of the
screening device of FIG. 3.
[0053] FIG. 5 shows a block diagram of the electrical components of
the screening device 30. The screening device 30 is based around a
microprocessor or central processor unit (CPU) 30 and the CCD 34.
The CCD 34 comprises an imager 82, and associated video digitiser
84 and video data interface 86. The screening device may also
includes a keypad 88 or may be operated via a combination of
buttons provided on the facia. The screening device also includes
electrically erasable read only memory (EEPROM) 90, dynamic random
access memory (RAM) 92 and the liquid crystal display (LCD) 56. The
EEPROM 90, RAM 92 and LCD 56 are connected to the CPU 80.
Alternatively, the EEPROM and RAM may be internal to the CPU. The
LCD 56 may be backlit and control is provided via a backlight
driver 94 which is connected to both the CPU 80 and the LCD 56.
[0054] The keypad 88 may be used to allow a user to enter data
required by the CPU 80 to control operation of the screening
device. Results from the screening device 30 are displayed to the
user via the LCD 56 which also acts to prompt the user for the data
required to operate the screening device. Power is supplied to the
CPU 80, LEDs 44, LEDs 58 and 60, microswitch 43 and CCD 34 from a
rechargeable battery pack 48. The batteries can be recharged from
the mains supply or, for example from a car cigarette lighter, via
an adaptor. The operation or recharging the batteries can be
controlled by the CPU or alternatively can be controlled manually.
Preferably, the screening device automatically shuts down to
preserve battery life if no cartridge is present or if the results
of the previous screening have been displayed for longer than a
preset time, say 5 minutes. Preferably, if an external power supply
is detected by the screening device the CPU 80 automatically
commences a battery recharging program. Preferably, the batteries
can hold enough charge to operate continuously for up to 24 hours
without being recharged.
[0055] The CPU 80 controls an electroluminescent backlight driver
94 to backlight the LCD 56. Preferably, the LCD 56 is capable of
displaying two rows each of 8 alphanumeric characters. In addition
to the LCD display, two LEDs, one red 58 and one green 60, are
provided. Illumination of the LEDs 56 and 60 is controlled by the
CPU 80 and may be used is to indicate visually the progress and
status of the scan, ie in progress, results ready for display, or
the outcome of the test. Alternatively, the progress or results
could be indicated by an audible signal. The LCD 56 may also
display status information.
[0056] In the embodiment described above the overall size of the
device is approximately 85 mm by 80 mm by 65 mm and the device
weighs approximately 300 g. The CCD 34 may be, for example, a
Connectix Quickcam, incorporating a CCD imager, video digitiser and
video data interface.
[0057] Operation of the screening device will now be described.
Disposable saliva test swabs 70 are stored in a sealed pack and one
swab removed immediately prior to use. The swab should be removed
from the pack by the person whose saliva is to be tested and is
wiped under the tongue for approximately 15 seconds. The swab 70 is
then inserted into the swab holder 16 of a disposable test
cartridge 10. Ten drops of a run fluid, which may be of any
conventional type, are added to the swab holder 16. The run fluid
transports the sample of saliva from the test swab 70 to the
absorbent pad 26 and onto the conjugate release pad 27, where the
saliva and run fluid mixture mixes with the labelled (e.g. with
gold, coloured latex particles carbon particles, fluorescents, or
any other suitable label) anti-drug antibodies. The sample
subsequently travels along the length of the nitro-cellulose
membrane 24. At each test zone any unbound labelled drug antibodies
are bound to the drug-protein derivative of the test zone. Any of
the labelled antibodies which have not been bound to the test zones
passes over the control zone where it becomes bound to the control
zone. The result is a number of lines of varying intensity spanning
the width of the membrane at points along the length of the
nitro-cellulose membrane corresponding to the drug-protein
derivative zones and the control zone. Each drug-protein derivative
zone can be used to detect a different drug. The higher the
concentration of the particular drug in the saliva sample, the less
intense the colour in that drug-protein derivative zone.
[0058] As soon as the test swab 70 has been located in the swab
holder 16 the test cartridge 10 is inserted into the screening
device by gently pushing the end of the cartridge furthest from the
swab holder 16 into the opening of the screening device 30,
allowing the test cartridge 10 to be guided by the receiving
bracket 32. The test cartridge 10 should be inserted gently until
the end of the test cartridge furthest from the swab holder 16
reaches the back 38 of the receiving bracket 32 when there will be
resistance against further insertion.
[0059] Once inserted into the screening device the cartridge is
left in position until the scanning process has been completed. A
message on the LCD and/or flashing of the LEDs indicates that the
scan is complete. Only then may the cartridge be removed.
[0060] As the test cartridge 10 is pushed into position it
displaces the micro switch 43. A signal is sent from the
microswitch 43 to the CPU 80 which activates the scanning process
by down-loading a preset program from EEPROM 90. Timer means are
provided to delay illumination of the immunoassay test until the
test has had time to run. Once the presence of a test has been
detected the CPU 80 commences initialisation by prompting the user
to set a timer to alert the operator to wait a sufficient time for
the sample to travel the length of the membrane. Alternatively the
user may time the test manually and an on/off power switch can be
provided which the user can operate once the test has been run and
the test cartridge 10 has been inserted into the screening device
30. The timer function may be provided by a separate timer
integrated circuit controlled by the CPU 80 or may alternatively be
provided internally to the CPU 80. When the prerequisite length of
time has elapsed, which is generally of the order of five minutes,
the timer sends a signal to the CPU 80 which alerts the operator
that the sample is ready for screening for example by flashing LEDs
58 and 60, displaying a message on LCD 56 or sounding an alarm. The
screening device is also able to time the test, analyse results,
output results and store the results automatically.
[0061] A plurality of adjacent membranes may be incorporated into a
single test cartridge with the membranes running longitudinally the
entire length of the cartridge from the absorbent pad to the end of
the cartridge furthest from the swab holder and each membrane 24
having a transverse width less than that of the test cartridge 10
such that a plurality of membranes, for example two, may be placed
side by side in the test cartridge 10. Processing the results of
the saliva test depends on identifying the intensity of the lines
on each membrane and relating each line to the drug which is the
subject of that particular test. Details of the number of adjacent
membranes in the particular cartridge and the number, type and
position of the drug-protein derivative zones and control zone on
each membrane are required for processing of the results. These
details can be held in the EEPROM and accessed by the CPU 80 upon
detection and recognition of the cartridge type or the user can
directly enter data required for the CPU 80 to recognise the test
cartridge 10. If the test cartridge 10 is to be recognised by the
CPU 80 it may carry appropriate marking such as a bar code, which
is read by appropriate means provided in the screening device and
the information is passed to the CPU 80.
[0062] The test cartridge 10 may be printed with the name of the
test which may be automatically read and identified by the CPU 80.
The test cartridge 10 may also contain an implanted microelectronic
circuit which may be interrogated by the CPU 80 by means of
electrical, infra-red or inductive links in order to ascertain
whether the cartridge is acceptable and to determine the nature of
the test.
[0063] The CPU 80 enables the CCD 34 and switches on the
appropriate LED 44, thus illuminating the window 22 of the test
cartridge 10. In the presently preferred embodiment three LEDs are
provided, with wavelengths of 430 nm, 565 nm and 660 nm
respectively. The wavelength of light emitted by each LED is chosen
with reference to the characteristics of the label, in particular
to its colour. Preferably the wavelength of the light used to
illuminate the immunoassay strip is complementary to the wavelength
of the particular label in order to provide the best contrast. In
the presently preferred embodiment, colloidal gold is employed as
the label and as colloidal gold is pink in colour a green LED with
a wavelength of 565 nm is used. Whilst the label has been described
as visible and the example of colloidal gold as a label would be
visible to the human eye, the label may be chosen to be visible to
the CCD array under certain lighting conditions and may not, either
under normal lighting conditions or under special lighting
conditions, be visible to the human eye.
[0064] Light from the LED 44 shines onto the window 22 of the test
cartridge 10 illuminating the nitro-cellulose membrane 24 visible
through the window 22. The illuminated membrane 24 is reflected by
the mirror 40 into the field of view of CCD 34. The image is
digitised and outputted via a video data interface to the CPU 80
for data processing. Preferably, the digital data is stored to
dynamic RAM for subsequent processing.
[0065] The image captured by the CCD 34 is skewed by the reflection
from the mirror 40 and the CPU 80 must first apply an algorithm to
correctly align the digitised data for processing. Preferably, the
CPU 80 runs an initial is correction algorithm to arrange the data
for subsequent processing. Preferably the initial correction
algorithm is set when the device is manufactured and, if necessary,
calibrated, at points during the life of the device rather than at
the beginning of each test.
[0066] Each cartridge may be used to run tests for a number of
different drugs. This can be achieved either by using a single
membrane with a larger number of drug-protein derivative zones or
using a number of membranes in a single cartridge. Up to eight or
more drugs may be analysed at any one time using a combination of
these methods. In the presently preferred embodiment, drug-protein
derivatives for cannabis (THC), cocaine (COC), opiates (OPI),
methadone, ecstasy and amphetamines (XTC) and benzodiazepines (BZO)
are bound to the nitro-cellulose membrane at discrete intervals.
The results for each of these drugs tests is indicated separately
by the screening device. Two panel tests, for example for methadone
and opiates, may also be provided. The data must then be segmented
such that each segment relates to one membrane only. The separate
segments are then processed separately. In the presently preferred
embodiment the CCD array is a Texas TC 255 P CCD array which is
made up of 324.times.240 elements. The digital data must be
segmented to correspond to the 344.times.240/N pixels covering that
particular membrane only where N is the number of membranes in the
cartridge.
[0067] Each membrane is therefore represented by an array of
p.times.q pixels where the p pixels span the length of the membrane
and the q pixels span the width of the membrane. The drug-protein
derivatives are bonded across the entire width of the membrane at
discrete intervals along the membrane. At any location (p,r) where
p falls within a particular drug-protein derivative zone the
intensity of the pixel is related to the amount of that particular
drug in the sample, regardless of the value of r in the range
0.ltoreq.r.ltoreq.q. The intensities of the pixels at (p,r) are
therefore summed over the range 0.ltoreq.r.ltoreq.q for each p.
[0068] Slight discrepancies between the theoretical position of the
membrane and the actual position of the membrane may be
accommodated by the screening device. The CPU 80 compares the
summed intensity at a specific location corresponding to the
theoretical centre position of the control zone with the intensity
at a predetermined number of adjacent locations to determine
whether there is any discrepancy between the theoretical location
of the control zone with the actual location of the control zone.
The CPU 80 applies a corresponding offset to subsequent
calculations if the theoretical and actual locations of the centre
of the control zone differ. The offset must be determined by
reference to the control zone because if any of the tests are
positive then the intensity of that drug-derivative zone will be
correspondingly reduced.
[0069] Alternatively, or in addition, the test strip 23 and test
cartridge 10 may be of contrasting colours. The unskewed data may
be processed using the contrast between the test cartridge 10 and
the test strip 23 to determine the actual location of the centre of
the test strip which may then be used to apply an offset to the
data if required.
[0070] FIG. 6 shows a typical graph of the resulting pixel
intensity against the location of the pixel for a single
protein-drug derivative zone and a single control, or reference,
zone. Once any offset of the membrane from the theoretical position
has been identified, the data is segmented according to whether it
lies in a drug-protein derivative zone, the control zone or in a
space between adjacent zones as shown in FIG. 6. Preferably, the
CPU 80 is a Hitachi H8/3002 microprocessor chip but any other
suitable microprocessor chip may be used. The CPU 80 segments the
data into a first plurality of data corresponding to the control
zone, a second plurality of data corresponding to the test zones,
and a third plurality of data corresponding to the background
zones. The CPU 80 then processes the first, second and third
pluralities of data, performing the following calculations to
determine whether each drug is present in the sample.
[0071] Define RW1 = p1 p2 .times. .times. RB = p2 p3 .times.
.times. RW2 = p3 p4 .times. .times. and ##EQU1## TW1 = p5 p6
.times. .times. TB = p6 p7 .times. .times. TW2 = p7 p8
##EQU1.2##
[0072] Then estimate REF = 1 - ( 2 .times. RB ( P3 - P2 + 1 )
.times. ( RW1 + RW2 ) ) ##EQU2## and ##EQU2.2## TEST = 1 - ( 2
.times. TB ( P7 - P6 + 1 ) .times. ( TW1 + TW2 ) ) ##EQU2.3##
[0073] If REF.ltoreq.0 then the reference, or control, zone has not
bound any of the products present in the saliva sample and run
fluid after it passed over the drug-protein derivative zones.
Either the control zone is faulty on the membrane or the test has
not been completed correctly which may be due to an insufficient
amount of run-fluid being added to the swab holder. The screening
device will display an error message and the cartridge should be
removed, reinserted and reread or disposed of and another cartridge
run. However, the delay for the test to be performed is not
required in these circumstances and the operator is provided with a
means for bypassing the timer operation to commence immediate image
acquisition and data processing. If an error is still detected then
the test must be re-run using a new cartridge and saliva
sample.
[0074] If REF>0 showing that the test has been successfully
completed but TEST.ltoreq.0 then the drug concentration in the
sample is such that all the antibody-gold conjugates have been
bound to the drug in the sample. The results of that test is set to
100%. The test is assigned a qualitative level "Positive". A
quantitative value would be represented as "greater than" a certain
level.
[0075] If TEST>0 and REF>0 then the test band concentration
is determined as follows: TEST .times. .times. BAND .times. .times.
CONCENTRATION = 1 - ( TEST REF ) ##EQU3##
[0076] The percentage of drug present in the sample is given by
100.times. Test Band concentration %.
[0077] The results for the concentration of each drug can be
displayed in a number of ways. The LCD 56 may be used to display
the name of the drug and its result. Alternatively only the fact
that the test for that particular drug is positive may be
displayed. If the display is to indicate a positive or negative
result only then the CPU 80 must have access to a threshold for
each drug which could be held in the EEPROM. For each drug if the
detected concentration exceeds the threshold then the result would
be positive and if the detected concentration falls below the
threshold then the result would be negative. Each separate
drug-protein zone must be tested in this way with reference to the
control zone to determine the concentration of that drug in the
saliva sample.
[0078] Alternatively, or in addition, positive and negative results
could be displayed using combinations of the LEDs 52 and 54
provided on the top of the screening device. In the presently
preferred embodiment, the red LED 52 will be continuously
illuminated and the green LED 54 intermittently illuminated to
indicate that the particular drug test is positive, and the green
LED 54 will be continuously illuminated with the red LED 52
flashing if the drugs test is entirely negative. The operator may
step through the result of each individual drug test by operating
the buttons 58 and 60 on the front of the screening device or may
view all the results simultaneously by down loading the results to
a pc with the necessary graphics facilities. Results may be stored
within the screening device until they are down-loaded to a PC. The
test image may be stored for subsequent downloading to a PC.
[0079] If the test indicates that any of the drugs are present in
the sample, follow up testing using an alternative test method may
be performed.
[0080] The CPU 80 is provided with a programming interface 98 to
allow the screening device to be programmed for example from a
remote PC. Serial and parallel PC links 96 and/or an infra-red link
may provided from the CPU 80 allowing the control of the screening
device to be relinquished to a pc or mainframe computer. Results of
the testing can also be displayed by the pc having been down loaded
from the screening device. Preferably, the CPU 80 is capable of
running self-testing diagnostic routines stored in EEPROM at
intervals which may be controlled either by presets in the CPU or
may be initiated on demand by the user.
[0081] In certain situations it may be preferable for the screening
device operator to be provided with a display indicating the image
produced by the CCD 34. An interface for the CCD 34 may be provided
to allow the operator to view the image on a small graphics
panel.
[0082] It may also be preferable to provide means for storing the
image of the person who provided the test sample. For this purpose,
the mirror 40 could be adjusted between the test screening position
and a second position which allowed the image of the person being
tested to be reflected onto the imaging means for storage and
subsequent retrieval. Additional optical apparatus, for example a
lens, may be required to modify the focal length along the external
light path.
[0083] FIG. 7 shows a block diagram of the electrical controls and
electrical apparatus which may also be used in the screening
device. Apparatus and controls which correspond to those of FIG. 4
are given the same reference numerals and reference should be made
to the description above.
[0084] In particular, a CMOS image sensor 82' may be used instead
of a CCD image sensor. A driver is associated with the CMOS image
sensor 82' and interfaces between the CPU 80 and CMOS image sensor
34'. A video buffer 86' replaces the video data interface 86 of
FIG. 5. Preferably, a Vision VV5404 imaging device having a
resolution of 356.times.292 pixels is used.
[0085] The electroluminescent backlight driver 94 shown in FIG. 5
may also be replaced by light emitting diode backlight driver 94'.
Furthermore, the volatile memory device 90 provided by an EEPROM in
the apparatus of FIG. 5 may be replaced by FLASH memory and/or the
non-volatile memory provided by the dynamic RAM (DRAM) in FIG. 5
may be replaced by static RAM (SRAM). An LCD 56' with a higher
resolution capable of handling graphics of 100.times.64 pixels may
also replace the LCD 56 of FIG. 5. This would allow all the test
results to be displayed simultaneously if required. Batteries which
are capable of holding sufficient charge to power the screening
device for up to 20 days may be provided.
[0086] Preferably, means 45 for adjusting the intensity of each of
the LEDs 44 may be provided. Adjustable current LED drivers may be
used as shown in FIG. 7.
[0087] The half silvered mirror 40 of FIGS. 3 and 4 may be replaced
by a plain first surface mirror 40. The angle of the mirror and the
imaging device may be altered to reduce the skew of the image. For
example, by adjusting the angle of the mirror 40 to 55.degree. to
the vertical and using a CMOS image sensor inclined at 10.degree.
to the vertical, the combined inclination of the CMOS image sensor
82' and the mirror 40 minimises the difference in the image width
over the width of the test strip with the result that the image
captured by the CMOS image sensor 82' is substantially unskewed
relative to the actual immunoassay test strip 23. In the case that
the image captured by the imaging device is unskewed, the CPU 80 is
not required to apply an algorithm to correct the digitised data
prior to processing the data.
[0088] Different numbers of LEDs 44 may be used to illuminate the
test strip 23. For example, four LEDs rather than three may be
mounted in a horizontal row parallel to the longitudinal length of
the test cartridge. If four LEDs are used, the two outermost LEDs
may be chosen to emit light of one wavelength whilst the two
innermost LEDs may be chosen to emit light of a different
wavelength. With this configuration, only one pair of LEDs may be
used to illuminate the immunoassay test strip 23 for the purpose of
determining the drug concentration. The remaining pair of LEDs may
be used for non-disruptive messaging, for example reading a bar
code on the test strip or cartridge. The intensities of the LED
pairs may be matched to provide optimal illumination of the
immunoassay test strip 23. Suitable wavelengths for the LED pairs
are 566 nm and 639 nm. However, the primary requirement in choosing
suitable LEDs is that the wavelength of light emitted is compatible
with the marker used on the immunoassay test strip 23 and that
illumination of any messaging markings does not corrupt the test
results.
[0089] The size and weight of the screening device may be affected
by the choice of electrical apparatus and controls. Using a Vision
VV5404, the overall dimensions and weight of the screening device
may be 210.times.70.times.50 mm and approximately 240 g.
[0090] The control band may only be used to verify that the test
has run successfully and may not be used for the quantification of
individual drug concentration calculations. In this case, null data
may be provided in order to quantify the test results. Such null
data may, for example, correspond to the data which would be
generated by illuminating a blank immunoassay test strip under
identical conditions to the illumination of the experimental
immunoassay test strip. Such null data would then give an estimate
of the intensity observed when the concentration of a drug in the
sample under test approximates or exceeds the amount of conjugated
antibodies released from the relevant pad. Such null data may be
compared to the test data to determine the concentration of the
substance in the sample under test.
[0091] Null data may be approximated by suitable filtering of the
experimental data eliminating any need for separate illumination of
an unused, clean test strip as a reference strip. For example, data
corresponding to the length and width of one of the background
zones may be interpolated to produce an estimate of the intensity
that is representative of null data. Prior to interpolation, the
data may be smoothed to improve the null data. More sophisticated
filtering techniques, including adaptive filtering, may also be
used in estimating the null data. Once null data has been estimated
or provided, the test data and null data may therefore be compared
to determine the concentration of the substances in the test
zones.
[0092] FIG. 8 shows schematically typical results of the appearance
of a control zone and test zone thus detected by a CCD or CMOS
image sensor on a test run on a sample of body fluid. The depth of
colour of the control and test zones vary over the width and length
of the zones resulting in an uneven appearance. However, in general
the depth of colour towards the Centre of the zones is deeper than
at the outer edges. When the test strip is illuminated by an LED of
complimentary wavelengths to the label used, the irregularities of
the test and reference zones result in higher optical absorbency at
the centre of the zones. Additionally, if the illumination over the
length and width of the strip is not uniform, the CCD or CMOS image
sensor will detect variations in the reflected lights intensity
which are entirely independent from the test results.
[0093] In order to reduce illumination irregularities and hence
suppress spurious test results, multiple error LEDs may be used to
illuminate the test strip. Preferably each LED has an individually
adjustable current. CPU 80 may be used to control the current
supply to each LED to reduce such illumination irregularities and
hence improve test results.
[0094] A further cause of the irregular appearance of the zones as
detected by the CCD or CMOS image sensor is the variation in path
lengths travelled by photons reflected from different parts of the
surface or the immunoassay strip. This effect cannot be eliminated
by a fixed pattern correction algorithm because to be most
effective such correction should take account of any slight change
of location of the test strip in relation to the mirror and/or CCD
or CMOS sensor. Differences, however small, between screening
devices mean that it is not possible to define a single fixed
pattern correction algorithm for all devices.
[0095] In order to minimise the effects of these variations the CPU
80 may digitally filter the data once the alignment algorithm has
been applied. Data corresponding to the entire test strip including
the control zone, test zones and background zones is filtered. Each
membrane is represented by an array of P.times.Q pixels where P
pixels span the length of the membrane (rows) and Q pixels span the
width of the membrane (columns). Across the width of the membrane
the intensity of pixels in each column would, under ideal
conditions, be identical. In practice, due to one or more of the
irregularities described, the intensity of the pixels in each
column vary to a greater or lesser degree. Thus the intensity of
the pixels in each column are summed and the mean value stored in a
1-d column intensity data array.
[0096] Next, the centre of each of the test, control and background
zones is estimated. The amount of marker deposited during the test
tends to be greater at the centre of the test and control zones
than at the edges of the zones. Hence, when the strip is
illuminated by an LED of a complimentary wavelength to the marker,
the intensity of the pixels reaches a local minimum close to or at
the centre of each of the test and control zones. The geometry of
the test strip being known, it is a simple matter to determine the
number of pixels spanned by any one zone. In practice, the test and
control zones preferably span 40 pixels. However, zones of widths
corresponding to any number of pixels reasonable to achieve the
desired resolution may be used.
[0097] The total intensity for each test and control zone is
estimated by summing the data in the 1-d column intensity data
array over that zone having used the local minimum intensity to
locate the centre of that zone and knowledge of the width of the
zone to determine how many data entries centred around the local
minimum entry correspond to that particular zone.
[0098] Although the method described so far takes into account any
irregularities in the deposit of the marker over the width and
length of the strip, it does not take into account any illumination
or uniformity. This may be achieved using information from the
irregularities detected in the background zones where no marker
should be deposited.
[0099] The data of the 1-d column intensity data array may be
corrupted by noise and noise reduction is therefore performed. The
filter achieves noise reduction by minimising excessive differences
between entries whilst retaining the underlying signal (intensity
of contrast between test zones and background zones). Any MA filter
producing a symmetrical response (ie one where no spatial
displacement or phase-shift is present in the output) may be used.
In the presently preferred embodiment, a moving average filter of
window length 3 columns is applied to the array. Longer window
length filters may produce better smoothing of the data and hence
improved noise reduction. The length of the filter window is
limited by the spatial distribution of the coloured particles
deposited. The window size of the filter must be chosen to be much
smaller than the spatial distribution of the coloured particles
deposited. Although longer window length filters produce better
smoothing, they require a large amount of memory for processing and
implementation is algorithmically less efficient than shorter
window length filters. However, the smoothing effect achieved by a
longer window length filter may be approximated by applying a
smaller window length filter multiple times to the data. The 1-d
data array is therefore copied and filtered three times using a
moving average filter of window size 3 columns. The output of the
filtering operation is retained in memory for use later as the
non-interpolated image data.
[0100] A second moving average (MA) filter is used to interpolate
the data to estimate a white, background level (ie to approximate
the data that would have been obtained had the test bands not been
present). The second filter must have a window length sufficient to
span the width of the control and test zones. It is applied to the
noise reduced data array which is outputted from the first MA
filter. Preferably, when the width of the test and control zones is
40 columns, the width of the second MA filter is 41 columns.
Preferably, the second MA filter is repeatedly applied to the
array. In one embodiment of the present invention, the second MA
filter is applied 21 times to the array. In particular, applying
the second filter a number of times to the data improves the
interpolation accuracy of the data corresponding to the test zones.
Applying the filter 21 times has been found to be particularly
effective for test zones which span approximately 13 elements of
the data array. The choice of window length is a compromise between
a requirement for excessive processing and achieving a reasonable
estimate of the white background level. The filter should
preferably be applied a number of times equal to or larger than
approximately 1.5 times the width of the test zones in the data
array entries. On each pass of the second MA filter each value of
the data array is only updated by the filter output if the filter
output is greater than the current value of that value of the
array. If the output of the filter is smaller than the current
value of the array, the current value is retained. This process
allows the effect of the absorbency across the test on control
zones to be minimised and a cleaner "white" background signal to be
estimated. Typically, when the white background level is estimated
from the 1-d data array (rather than on the original 2-d data
array) the performance of the above described process is superior
to alternative methods using interpolation of the background by
applying a polynomial curve fitting approach. The output of the
second filter is supplied to a third moving average filter.
[0101] The third MA filter is applied to the output of the second
MA filter to remove any spurious peak values or spikes in the data.
The third MA filter of width 11 columns is preferably applied twice
to the array. Preferably, the window length of the third MA filter
is chosen to be approximately equal to the width of the test zones
in pixels. When the width of the test zones is approximately 13
pixels as in one presently preferred embodiment, a choice of 11 for
the window length of the third MA filter is presently preferred.
The processed, white background array is then used with the total
intensity data generated for each test and control zone to estimate
the ratio of intensity in each test zone for the background zones
and the intensity of the control zone to the background zone. The
resulting ratio for the control zone is then compared against a
pre-set threshold to confirm that an appropriate amount of the
marker has been deposited in the control zone to indicate that the
sample has run successfully and that the amount of marker deposited
in the control zone corresponds within limits to the amount
expected. If the results of the comparison are negative, a fault
condition is reported to the user, the results are ignored and a
repeat test is required.
[0102] It will be readily apparent to a person skilled in the art
that the filtering operation could be performed differently to
achieve the same effect. For example, the white background signal
may be estimated from the 2-d data array by using linear estimation
of several pixel pairs located at offsets approximately half the
inter-test zone spacing from the central pixel whose background
value is being interpolated. Typically, these pairs lie in the
white background zones between test zones and their mean value
gives a good estimate for the background intensity in test zone
locations. Interpolation of the 2-d data array may be carried out
and column totals taken subsequently to form the 1-d data array. An
advantage of this technique is that the location of the window of
the cartridge (and therefore of the relevant portion of the test)
may be performed more reliably. The test zones are virtually
eliminated from the 2-d array without introducing other distortions
allowing the periphery of the window of the cartridge to be located
more easily. This method may be particularly effective at
accurately interpolating the background when the test zones are
very faint. Faint test zone colour concentration frequently arises
for drug tests where the concentration of drug present in the
sample is close to the threshold value which determines whether the
test is positive or negative.
[0103] For use in adverse weather conditions, various adaptations
(not shown in FIG. 1) to the test cartridge may be provided. A
retractable, transparent cover may be provided on the test
cartridge to protect the immunoassay test strip 23 which is
otherwise exposed through the window 22, for example from exposure
to rain. The window is retracted automatically upon insertion of
the test cartridge into the screening device and is redeployed when
the test cartridge is removed from the screening device. As shown
in FIG. 9, the run fluid may be contained in a trough 68 within the
cylindrical swab holder 16 of the test cartridge 10. The run fluid
is held in place by a thin penetrable membrane 66 that covers the
trough 68 until the test cartridge is used. The membrane 66 is
pierced by spike 64 when the membrane 66 is deformed downwardly by
the introduction of a test swab 70 into the swab holder 16. The run
fluid is drawn by capillary action across to the conjugate release
pad 27 via the saliva collection pad 72 of the test swab 70. If the
operator exerts too much force on inserting the test swab 70 into
the swab holder 16, the membrane 66 may rupture in an explosive
manner causing the run fluid to be splashed onto the conjugate
release pad 27 without first passing over the saliva collection pad
72. If the operator does not use enough force on inserting the test
swab 70 into the swab holder 16, the membrane may not be pierced by
the spike 64 and the test will not run. These problems are
minimised and operator error reduced or eliminated by providing an
internal thread on the bore of the swab holder 16 and the handle 74
of the test swab 70 may be provided with an external thread. The
saliva collection pad 72 is placed into the swab holder 16 and the
handle 74 of the test swab 70 is screwed into the swabholder 16
until it reaches an end atop. This allows controlled piercing of
the membrane 66 on trough 64 and hence the controlled release of
the run fluid to the saliva collection pad 72.
[0104] A second, elastic membrane with an aperture may be
positioned above the run fluid membrane in the cylindrical swab
holder 16. The aperture of the elastic membrane expands to allow a
test swab to be inserted through the aperture and would form a
waterproof seal around the test swab 70 prior to the test swab
piercing the run fluid membrane. Upon removal of the test swab 70,
the aperture of the elastic membrane contracts preventing fluid,
other than that on the test swab, from entering the test
cartridge.
[0105] FIG. 10 shows a cut away side view of a second embodiment of
a test swab which incorporates both the spike 64 and the run fluid
within the test swab, and thus removes the need for these to be
provided in the test cartridge.
[0106] The handle 74 of test swab 70 is made from a transparent
outer tube 78 open at the lower end thereof and closed at the top
end thereof. At the lower end of the outer tube 78 a flange 76 is
provided which protrudes outwardly from the outer tube 78. The
flange 76 may extend around the entire periphery of outer tube 78
or may extend only around part or parts of the periphery of outer
tube 78. An adequate spike 64 extends from the closed top end of
the outer tube 78 downwardly. Alternatively, instead of a spike, a
pin or any other sharp protrusion may be used.
[0107] A saliva collection pad 72 is attached to the lower end of
tube 79. The tube 79 is open at both ends. The diameter of the bore
of the tube 79 is large enough to allow the spike 64 attached to
the outer tube 78 to enter the bore. Adjacent to the saliva
collection pad 72 inside the bore of tube 79 are positioned in
order a filler pad 101, dye release pad 102, a dye receptor pad 100
and a run fluid capsule 68'. In the embodiment shown in FIG. 10,
the run fluid chamber 68' is a capsule. The run fluid capsule 68'
is positioned closest to the upper end of the inner tube 78. The
tube 79 has a diameter slightly less than the internal diameter of
the outer tube 78 such that the tube 79 may be positioned within
the outer tube 78 and moved vertically relative to the outer tube
7a. To assemble the swab, a s spring 77 is placed in the outer tube
78 through the open lower end and the tube 79, which effectively
forms an inner tube, is inserted into the outer tube 78 to hold
captive the spring 77. In the assembled position, the saliva pad 72
and lower part of the tube 79 encasing the filter pad 101, dye
release pad 102, dye receptor pad 100 and run fluid capsule 68'
protrude below the lower end of the outer tube 78. The spike 64 is
held remote from the capsule 68'. In this sampling position, the
swab may be used to collect a sample of bodily fluids. The length
of the spike is determined by the maximum displacement of the tube
79 relative to the outer tube 78 such that the spike 64 may be used
to puncture the run fluid capsule 68' when the tube 79 is forced,
against the pressure of the spring 77, into the outer tube 78. The
tube 79 is transparent and the outer tube 78 may also be
transparent.
[0108] With the elongate spike held in the sampling position (i.e.
disbursed from the capsule 68'), a saliva collection pad 72 of the
test swab 70 is inserted into the mouth of the person to be tested
and saliva collected until the saliva migrates by wicking effect up
the saliva collection pad 72 through the filter pad 101, through
dye release pad 102 where it becomes coloured by the dye and into
the dye receptor pad 100. Once the dyed saliva becomes visible on
the dye receptor pad 100, the user is alerted that an adequate
sample of saliva has been collected. The user then takes the test
swab 70 and inserts it into the swab holder 16 of the test
cartridge 10. As shown in FIG. 11, once the saliva collection pad
72 of the test swab 70 contacts the conjugate release pad 27 of the
test cartridge, the tube 79 is prevented from being further
inserted. Continued pressure on the outer tube 78 of the test swab
70 causes the outer tube 78 to move downwardly relative to the
stationary tube 79 against the pressure of the spring 77 moving the
elongate spike from the sampling position to the sample
transferring position. As the outer tube 78 is depressed relative
to the tube 79, the spike 64 moves downwardly in the bore of the
tube 79 and pierces the capsule 68' causing the run fluid to move
under gravitational force and capillary action through the dye
release pad 102, dye receptor pad 100, filter pad 101 and onto the
saliva collection pad 72 where it mixes with the saliva. It is then
drawn onto the conjugate release pad 27 which is in intimate
contact with the saliva collection pad 72. The swab holder 16 is
provided at its upper end with resilient lips 104 which extend
inwardly from the cylindrical swab holder 16. The lips 104 define a
hole with a diameter which is slightly larger than that of the
outer tube 78 but smaller than the diameter of the flange 76 on
outer tube 78. The flange 76 is profiled such that there is a
smooth change in diameter of the flange at the open end of the
outer tube 78 and a more abrupt change at the upper extent of the
flange 76 remote from the open end of the outer tube 78. Upon
insertion of the outer tube 78 into the swab holder 16, the gradual
change of diameter of the flange 76 forces the resilient lips of
the swab holder apart allowing the outer tube 78 to pass into the
swab holder 16. Once the outer tube 78 has been inserted to a point
just beyond the extent of the flange 76, the lips of the swab
holder 16 spring back to their original unstressed position and the
outer tube 78 is effectively held in position relative to the swab
holder 16 of test cartridge 10 preventing the test swab 70 from
being accidentally removed from cartridge 10 and hence ensuring
intimate contact of the saliva collection pad 72 and the conjugate
release pad 27 for the duration of the test. The filter pad 101 may
be used to prevent the dye from compromising the test results.
Alternatively, a dye may be chosen which, although visible to the
human eye, is invisible or nearly so, at the LED wavelength used by
the screening device. The filter pad 101 may also limit the rate at
which the run buffer is released from the pierced capsule 68' to
the test cartridge 10, thereby improving the mixing of the saliva
sample and the run fluid.
[0109] FIG. 12 shows a cut away side view of a third embodiment of
a test swab which is suitable for use with a test cartridge. This
embodiment of test swab 70 requires that a spike 64 is provided in
the swab holder 16 of the test cartridge. As shown in FIG. 14 the
spike 64 is attached to a spike holder 65 which in turn is attached
to the swab holder 16. The spike 64 preferably has a cruciform
cross-section as shown in FIG. 15. The test swab 70 comprises a
saliva collection pad 72, a main tube 108, a run fluid chamber, or
capsule, 68' and an indicator section. The indicator section
comprises a capillary tube 110, a dye release pad 102 and a dye
receptor pad 100. The saliva collection pad 72 is in communication
with the open base of a main tube 108. The upper end of the main
tube is also open. A penetrable gelatine capsule 68' filled with
run fluid is located within the main tube 108 spaced from the
saliva collection pad 72. Disposed to the side of the main tube 108
is a capillary tube 110. A small port 116 is provided in the wall
of the main tube 110 a short distance from the saliva collection
pad 72. The open lower end of the capillary tube 110 is attached to
the main tube 108 at the port 116 and communicates with the main
tube 108 via the port 116.
[0110] Provided around the periphery of the lower portion of the
main tube 108 and capillary tube 110 is a guide 112 and insertion
endstop 106. The end stop is spaced from the saliva collection pad
72 substantially the same distance as the distance from the centre
of the capsule 68' to the saliva collection pad 72. The guide 112
and end stop 106 may extend around the whole periphery of the main
and capillary tubes 108 and 110 or only partially around the
periphery.
[0111] At the end of the capillary tube 110 remote from the port
116, a dye release pad 102 and dye receptor pad 100 are provided.
The dye receptor pad 100 is positioned at the open end of the
capillary tube 110 and is visible from the top and/or sides of the
capillary tube 110. The dye release pad 102 is spaced from the dye
receptor pad 100. In operation, as saliva is collected on the
saliva collection pad 72, some of it is drawn up the capillary tube
110 by capillary action where it contacts the dye release pad 102.
The saliva becomes dyed as it passes over the dye release pad 102
and as it travels further up the capillary tube 110 it contacts the
dye receptor pad 100 which becomes visibly stained by the dye
indicating that an adequate sample of saliva has been collected.
The test swab 70 may then be removed from the mouth of the person
being tested and inserted into the test cartridge 10. The guide 112
ensures that the test swab 70 is inserted optimally into the swab
holder 16 and further ensures that the run fluid capsule 68' will
be pierced by the spike 64 of the swab holder 16. The end stop 106
of the test swab 70 prevents the user from inserting the test swab
70 too far into the test cartridge and thus prevents the test
cartridge from becoming damaged which may interfere with the proper
running of the test. The user is alerted by the test stop hitting
the periphery of the swab holder 16 that the test swab 70 is fully
inserted and has been advanced far enough into the swab holder 16
for the spike 64 to have punctured the capsule 68' thereby
releasing the run fluid which mixes with the saliva sample and is
transported by gravitational and capillary action onto the
conjugate release pad 27 of the test strip 23. The dye receptor pad
100 may be a cotton swab.
[0112] The guide 112 may be formed of simple projections with the
diameter of the swab measured across the projections being slightly
smaller than the diameter of the swab holder 16 such that when the
swab is inserted into the swab holder 16 the guide 112 causes the
swab to be centred in the swab holder 16. Alternatively the guide
112 may consist of an external thread arranged around the periphery
of the main and capillary tubes 108 and 110 and co-operating with
an internal thread provided on the bore of the swab holder 16. If
the guide is provided by an external threaded portion then the end
stop 106 may be omitted and the swab inserted until the end of the
thread is reached determining the final position and pressure of
the spike 64 on the capsule 68'. Other embodiments of the test swab
may be provided.
[0113] Although described with reference to lateral flow
immunoassay testing, the above described test swabs could be is
used to take a sample of saliva for agglutination testing. The
difference being that instead of the saliva/run fluid mixture being
drawn over a conjugate release pad and thereafter onto and along a
nitrocellulose test strip, an agglutination test cartridge is
provided.
[0114] The screening device may also be used to detect the results
of agglutination tests. Agglutination tests are generally
categorised into one of two categories depending on the size of the
analyte whose presence is to be detected. The screening device may
be used to determine the results of both agglutination
categories.
[0115] In the first category, large analytes with multiple epitopes
(binding sites) such as proteins can be detected. Coloured (or
white) latex beads (microspheres or nanospheres) are coated with
antibodies to the protein, suspended in an appropriate buffer
solution, mixed with the sample under test and the mixture
incubated. The presence of the antigen (ie the analyte whose
concentration or presence in the sample is being tested and to
which the antibodies are directed) in the sample causes multiple
latex beads to bind together by bridging between two antibodies
coated to different beads. Because the proteins (or other large
molecule under test) are capable of binding with more than one
antibody at a time due to their multiple epitopes and each latex
particle has multiple antibodies coated to it, then complexes of
beads are formed causing agglutination. Huge molecules, which are
discernible to the naked eye, are formed by the large scale
clumping. These molecules may be detected by the screening device
and the relative concentration of the substance under test may be
calculated. Use of the screening device allows the sensitivity of
the test to be increased because individual pairs of bound latex
beads (dimers) can be detected by using imaging apparatus with
sufficient optical magnification.
[0116] There are further advantages in using the screening device
for the second category of agglutination reactions. In this second
category, sometimes referred to as agglutination inhibition
reactions, smaller analytes such as drugs can be detected. Coloured
(or white) latex beads (or polystyrene beads or liposomes) which
are irreversibly attached to drug molecules are manufactured. Free
antibodies are mixed with the sample under test and then added to
the coloured latex beads. If there are no drug molecules present in
the sample, the antibodies bind with the coloured latex beads
forming bridges between beads and agglutination which results in
localised high concentrations of coloured latex beads which can
then be detected. However, free drug molecules in the sample will
compete for binding sites on the free antibodies with the drug
bound latex beads. Hence, if a sample contains a drug being tested
for, the drug molecules bind to the free antibodies which are not
then free to bind with the latex beads, hence presence of the drug
inhibits the agglutination which would otherwise occur. The
absence, or reduction, of agglutination can be detected and the
concentration of the drug in the sample may be calculated.
[0117] Other variants of these agglutination reactions such as
using the device to monitor the rate of agglutination or the use of
different types of particle or different reaction mechanisms will
be obvious to those familiar with this field.
[0118] An embodiment of the screening device may also be used to
screen agglutination reactions. In order to determine the results
of an agglutination test, regions of (bio) chemical coagulation or
agglutination (hereafter referred to as condensates) must be
uniquely identified, counted, measured in area, colour and/or
intensity and generally distinguished from one another. In the
digitised image, the condensates of interest are generally larger
than a single pixel and adjacent pixels belonging to the same
condensate must be recognised as such. Distinct condensates must be
differentiated. There are various sources of noise in the digitised
image. Fixed pattern and random (statistical) distortions are
introduced by the optical components. Spatial location and
variations due to manufacturing tolerances of the Components of the
screening device also introduce random errors. Compensation for
these errors is provided by the screening device. Correction for
non-uniform illumination and differing imaging parameters (such as
exposure and amplification) and variations in the concentrations of
the test sample and reaction chemistry may also be corrected.
Objects within the image which are too small, too large or of a
particular shape may be disregarded. In the preferred embodiment,
the instrument is hand held and powered by batteries. The image
processing is capable of producing results accurately and rapidly
using moderate computer processing and memory resources.
[0119] FIG. 21 shows the plan view of a single reaction
agglutination test cartridge. The test cartridge 10' is made of
three sandwiched plastic layers. The middle layer defines the sides
of the channel 230, the sides of the reaction chamber or chambers
232 and the sides of the overflow reservoir 234. The top layer
defines holes forming venting holes 236 to the overflow reservoir
234 when the layers are assembled. The top layer also defines an
entry port 16' into which the sample may be inserted. The entry
port 16' communicates with the channel 230 provided in the middle
layer. The bottom layer and top layer form the bottom and top
respectively of the channel 230, reaction chamber 232 and overflow
reservoir 234.
[0120] As described above, a sample of bodily fluid such as saliva
is pre-processed by mixing with free antibodies to the drug under
test and coloured latex beads which are irreversibly attached to
the drug molecules under test, and the mixture applied to the test
cartridge 10' using entry port 16'.
[0121] The sample is drawn from the entry port 16' along the
channel 230. The channel is designed to have a length which allows
time for any pre-processing reactions to occur. Once it has passed
the length of the channel 230, the sample enters the reaction
chamber 232 where agglutinates develop. An overflow reservoir 234
is provided in the test cartridge 10' and communicates with the
reaction chamber 232. Once the reaction chamber is filled by the
sample, excess sample moves to the overflow reservoir 234 which has
venting holes 236 which are exposed to the atmosphere. The excess
sample may therefore escape from the test cartridge 10' preventing
pressure build-up inside the cartridge.
[0122] A window is provided in the top of the reaction chamber 232
and the window is illuminated by the screening device to detect the
results of the agglutination reaction. Alternatively, the entire
test cartridge 10' may be made from a plastics which is transparent
to the wavelength of light used by the screening device.
[0123] FIG. 22 shows an agglutination test cartridge used to
evaluate the presence or absence of multiple analytes. In addition
to a plurality of reaction chambers 232, the test cartridge 10'
provides a plurality of reactant chambers 238. Each reactant
chamber 238 holds a supply of coloured latex beads irreversibly
bound to molecules of the drug for which the test is being
conducted. The reactants are held in an immobilised state for
example by freeze drying. The saliva sample to be tested is mixed
with free antibodies to the drug under test and supplied to the
test cartridge 10' via the entry port 16'. The saliva/free antibody
mix passes along the channels 230 into the reactant chambers 238
where it mixes with the coloured latex beads. The mixture then
passes to the corresponding reaction chamber 232 where any is
agglutination reaction occurs. By using coloured latex beads bound
to different drug molecules in each reactant chamber 238, the
presence of a number of different drugs can be detected using a
single test cartridge 16'. Each reaction chamber 232 communicates
with the overflow reservoir 234 which has venting holes 236 exposed
to the atmosphere. The reactant chambers 238 may be placed in
series or in parallel as required by the test. In the embodiment of
test cartridge 10' shown in FIG. 22, saliva/free antibody mixture
is supplied directly to two of the three reactant chambers 238 with
the third reactant chamber 238 being supplied with the mixture from
one of other reactant chambers.
[0124] Alternatively, the reactant chamber 238 could hold a sample
of free antibodies to the drug under test and the coloured latex
beads irreversibly bound to the drug under test could be mixed with
the saliva sample before the mixture is supplied to the entry port
16'. Three reaction chambers 232 are provided and the channel 230
bifurcates to allow entry of the sample to two of the three
chambers 232. The third chamber is supplied with sample directly
from one of the other chambers.
[0125] Windows are provided in the top of each of the reaction
chambers and when processing the data using an embodiment of the
screening device, preset data is provided to give the location of
the different reaction chambers.
[0126] The image processing for agglutination reactions will now be
described with reference to FIGS. 16-19.
[0127] FIG. 16 shows a flow chart of the screening operation. The
main difference between testing for an agglutination reaction and
for an immunoassay test is that the CPU processes the digitised
image in a different manner. The hardware of the screening device
may be identical for screening immunoassay tests and agglutination
reactions. Alternatively, the agglutination reaction may use a
different size of test cartridge and the opening in the screening
device may be adapted to accommodate the agglutination reaction
test cartridge. An adaptor may be provided to allow two or more
sizes or types of test cartridge to be used.
[0128] The image is acquired, digitised and the data stored to a
2-D array (120) by the imaging section of the screening device. The
data array requires around 102 KB of memory (356.times.292
pixels.times.8 bits per pixel). The data processing can be
conducted using a small amount of additional memory of say 8 KB
thus eliminating the need for providing memory for a second array.
The screening device is therefore provided with commonly available
128 KB static RAM, Fixed pattern image distortions are removed by
applying various two-dimensional transformations to the data (122).
These techniques can be used to remove tilt, trapezoidal, rotation,
translation, pin-cushion (where the appearance of the image is
distorted so that the centre of the image appears to have been
pulled upwardly out of the plane of the image) or barrel distortion
(where the appearance of the image is distorted so that the centre
of the image appears to have been pushed downwardly through the
plane of the image) of the image. Suitable 2-d transformations are
simple mathematical equations for mapping the points of the
distorted image to the corresponding points on the original object.
If a continuous transformation is determined, it must be
discretised for implementation in the processor. Alternatively, the
transformation which maps the original image onto the distorted
image may be estimated, in which case the inverse transformation
must be calculated for application in the processor.
[0129] For agglutination test, the number of small condensates can
be large and the accuracy of the result is limited by image
resolution and distortion. It is therefore desirable to transform
the image prior to any further processing to permit image scaling
and the removal of distortion to be consistently achieved. By
measuring for each image the location of the corners of the test
window and the mid-points of the test window edges, an algorithm
may be developed which calculates the mathematical transformation
necessary to achieve a target window size, position, rotation and
distortion.
[0130] Once any imaging distortions have been corrected, noise
reduction (124) is performed by applying a 2-D low pass filter to
the data. Once the data has been filtered, histograms of pixel
intensities for the entire array are created (126). If necessary,
histograms of pixel intensities of sub-sections of the array may be
created. Using the pixel intensity histograms and knowledge of the
underlying chemistry of the agglutination reaction, threshold
intensities for sub-sections of the image are produced (128). For
example if 20% of the image area is normally occupied by
condensates, then the histogram can be used to determine the pixel
intensity thresholds to divide the image sections into two
portions, one approximately four times as large as the other. This
information is test specific and is stored on preset data in the
memory of the screening device. Once the thresholds have been
determined, they are applied to the array to transform it into a
monochrome 2-d array (130). Depending upon the chemistry involved,
portions above (or below) the threshold value determined for each
image section are interpreted as agglutination condensates or
background regions. Condensate areas are then represented by
negative integer values and the background regions by a zero value.
Different image sub-sections may be represented by different
negative integer values.
[0131] The monochrome array is first processed to identify the
background regions (zero valued entries) and a plurality of
positively valued entries which closely approximate the location of
the condensates (132). Flow charts for processing the monochrome
array to identify background regions and condensates are shown in
FIGS. 17 and 18. FIG. 18 is a flow chart showing the processing of
the data during the scan. The image array address registers X and Y
are initialised to zero (160). A current block register is
initialised by setting the value to one (162). The monochrome array
is sequentially processed. The order in which the array is
processed is shown in FIG. 17.
[0132] FIG. 17 shows the sequence of the raster scan. Processing
commences with the data which corresponds with the bottom lefthand
corner of the image, works along one column of the agglutination
test, moves to the adjacent row and starts processing the entries
corresponding to the next column.
[0133] The elements are referenced by (X,Y) where
0.ltoreq.X.ltoreq.Xmax and 0.ltoreq.Y.ltoreq.Ymax with X
incrementing column-wise from left to right and Y incrementing
row-wise from bottom to top. Elements in the array which store
negative integers correspond to condensate areas.
[0134] As each element is accessed a check is made to determine
whether the value stored in that element is negative (164). If the
element is negative, the value of element is set to the value of
the current block (166). The value of the current block is
increased by one (168). The surrounding elements of the array which
have already been scanned are identified. For example FIG. 17 shows
as the current position element the central element. The adjacent
previously scanned elements are those numbered 12, 7, 8 and 9. The
values of these previously scanned adjacent elements are compared
with each other and with the value of the current element. The
smallest non-zero positive value of the current element and
previously scanned adjacent elements is stored (170). Assuming that
this value is stored as Z, the current block value is compared to
the value of Z+1(172). If the value of Z+1 is smaller than the
value of the current block all non-zero positive elements among the
previously scanned adjacent elements and the current element are
tagged (174), that is the value stored in the element is set to the
value Z, the current block is decreased by one (176) and the
processor moves on to consider the next element in the array (180,
182 and 184). If the value of Z+1 is larger than or equal to the
value of the current block then the current element pointer is
incremented by one (178) and the processor moves on to consider the
next element in the array. Processing continues until each element
of the array has been processed in this manner (180, 182, 184).
[0135] Some of the condensates within the array will have been
identified more than once by the above processing and areas of a
single condensate may have different values or "tags". Where more
than one tag relates to a single condensate, further processing to
resolve the discrepancy is required. Tags which are equivalent, ie
relate to the same condensate, must be identified and the
information used to amend the array such that the same tag is used
for each distinct condensate but different condensates are
identified by different tags. A 1-d "tag tree" array is constructed
(134). The length of the tag tree array is set by the value of the
final current block from processing the monochrome array. For
example, if the current block is say 5 after finishing the first
processing, the tag tree array is a (1.times.5) array. The tag tree
array is initialised such that each element references itself i.e.
array(element)=element. Each tag (condensate label) is therefore
mapped onto itself. By modifying the elements of the tag tree
array, it is possible to identify equivalent tags. To do so, the
processed monochrome array is processed a second time.
[0136] FIG. 19 shows a flow chart of the second processing
operation. The processed monochrome array is processed element-wise
in the same manner as described with reference to the first
processing. For each element, the values of the current element and
those previously processed array elements adjacent to the current
element are compared (192). If any positive, non-zero entries are
found, the lowest value is selected and stored in the tag tree
array element corresponding to the higher value tag or tags (196).
For example if the current element has a value 3, and the adjacent
elements which have already been processed have the values 2,3 and
4, then the value 2 is stored to the third and fourth tag tree
elements. The equivalence between tags is thus recorded.
[0137] The elements of the 1-d tag tree are scanned from the first
entry to the last and updated. FIG. 20 indicates the processing
carried out. A tag tree address register X is initialised by
setting all the entries to zero. X is valid over the range 0 to
Xmax where Xmax is equal to one plus the number of elements in the
tag tree array (210). The current tag tree entry is stored to a
storage register Y (212). The element of the tag tree array
corresponding to the value stored in storage register Y is accessed
and its value stored in register Z (214). The current tag tree
entry (X) is set to the value of register Z (216) and the X
register incremented to process the next tag tree element (218)
until all elements have been processed (220). For example for a 1-d
tag tree containing a thousand entries numbered 1 to 1000 the
initialised array is TREE[LEAF]=LEAF. After initial processing, it
is possible for some entries (leaves) to be equivalent to others.
For example, TREE[752=329, TREE[839]=329, TREE[329=78 and
TREE[78]=78. Entry 78 is equivalent to itself and is therefore a
primary leaf. The remaining entries of this example can be related
to the primary leaf by a branch [752,839]-[329]-[78] being the
branch in this example. Scanning the tag tree decomposes the
entries to leave all the entries
TREE[752]=TREE[839]=TREE[329]=TREE[78]=78. The entries of the tag
tree are then limited to primary leaf values thereby eliminating
branch values and setting all the entries of a branch to the same
value.
[0138] When the scan is completed, only legitimately distinct
"leaves" of the tree remain; the branches linking the leaves have
been eliminated. The leaves are not necessarily contiguously
arranged nor are they necessarily in ascending numerical order.
[0139] Referring now to FIG. 16, a tag usage array of identical
length to the tag tree array is initialised by setting all the
elements to zero (140). Each element of the updated tag tree is
checked to see whether it is non-zero. For each non-zero value of
the tag tree, the corresponding array element of the tag usage tree
is set to one or any non-zero value (142). The current block
register is then used to consecutively number the tag usage tree
values. The current block register is set to one. In an
element-wise fashion, each element of the tag usage tree is
compared to zero and each time a non-zero entry is found, its value
is set to the value of the current block register which is then
incremented (144).
[0140] The tag tree is then updated once more using the tag usage
array. The value of the current tag tree array element is used to
reference the corresponding tag usage tree element. The value of
the current tag tree array element is replaced by the value of the
referenced tag usage tree element (146).
[0141] Once the tag tree has been updated, the elements are
compared and the highest positive value indicates the number of
distinct condensates detected in the original image (148). The
processed monochrome array can be updated using the tag tree array
(150). The array is processed element-wise as before and where the
value stored for an element is positive and non-zero a check is
made of the tag tree array to determine whether or not the value
should be amended. Assuming the current element of the processed
monochrome array is N and the value stored therein is 3, the
3.sup.rd element of the tag tree array is accessed. The value
stored in the 3.sup.rd element of the tag tree array is then stored
at the Nth element of the processed monochrome array. The twice
processed monochrome array now represents the background regions
denoted by zero-valued entries and contiguously numbered distinct
condensates denoted by contiguous positive numbers. The condensates
may be classified according to their size and relative frequency by
counting the number of entries with the same positive non-zero
values in the twice processed monochrome array (152).
[0142] The results obtained are used to determine whether the
sample contained the analyte(s) being tested for (154). In the
simplest case, the agglutination test measures just one analyte.
For standard tests, high numbers of large agglutinates indicate a
positive result whereas for inhibited agglutination reactions, this
indicates a negative result. Depending on the properties of the
noise reduction filter (124) and threshold determination (128),
large numbers of small agglutinates may or may not be obtained.
Therefore, the agglutinates may have to be categorised according to
their size. This is achieved by performing a final image scan in
which the number of times a particular tag occurs is recorded in a
new data array. Agglutinates of size smaller than a given value are
indicated by the new array having a value less than the threshold.
These agglutinates may be disregarded in the results of the
test.
[0143] In a manner similar to that of processing the results of
agglutination tests, the screening device may be used to determine
the outcome of precipitation reactions, reactions based on
electrophoresis, immunoelectrophoresis, immunofixation
electrophoresis, enzyme immunoassay and immunofluorescence. Any of
these techniques may be adapted to allow the screening device to
differentiate between the presence and absence of analyte in a
patient sample by appropriate image processing. For certain
reactions, the test may be designed to produce a colour change
which can then be detected by the screening device. Kinematic
analysis may be used both to determine the rate of change in colour
and to determine the final test result. The screening device is
capable of performing many kinds of kinematic analyses. Combination
tests may be provided with the test bands for multiple analytes
co-located on the test strip by ensuring the optical absorbency of
each test is independent. The screening device may then
differentiate between the independent test bands by scanning first
with one optical wavelength and then with the next wavelength.
[0144] Channel-based reaction sequences may be designed such that
intermediate reaction products deposited on the nitro-cellulose
strip may be detected optically as the reaction progresses.
Intermediate reaction products may allow for a reliable early
warning of the test results before the test has been completed.
[0145] The latex beads may be bound to either analyte (drug),
antibody or both. Bonds can form between two analytes via an
intermediary, or carrier, molecule known as a protein bridge (for
example polylysine). Reactions may be organised into one of two
classes: competitive or non-competitive. These classes are akin to
the distinction between agglutination and inhibition of
agglutination reactions.
[0146] Kinematic analysis, where the rate of change in colour is
determined, may also be undertaken by the screening device. The
screening device may be provided with a plurality of illumination
sources of differing wavelengths. Combination tests can be designed
such that the test bands for multiple analytes are co-located but
the optical absorbency is independent such that the screening
device can differentiate between the results for the multiple
analytes by illuminating the test strip with different wavelengths
of light successively.
[0147] Instead of providing printed test zones spanning the entire
width of the nitro-cellulose strip, it is possible to print an
array of dots with each separate dot comprising a test zone for a
different analyte. Where there are only a small number of analytes
to be tested, the separate dots and the control band or dots may be
printed closer to the conjugate release pad to reduce the run time
of the test.
[0148] It is also possible that an assay test is designed whereby
intermediate reaction products are deposited during the reaction
process. These reaction products may be detectable prior to the
completion of the test and in certain circumstances, it is
desirable to provide an early output where the results of the test
can be reliably reported using the intermediate reaction
products.
[0149] With respect to the above description, it is to be realized
that equivalent apparatus and methods are deemed readily apparent
to one skilled in the art, and all equivalent apparatus and methods
to those illustrated in the drawings and described in the
specification are intended to be encompassed by the present
invention. Therefore, the foregoing is considered as illustrative
only of the principles of the invention. Further, since numerous
modifications and changes will readily occur to those to skilled in
the art, it is not desired to limit the invention to the exact
construction and operation shown and described, and accordingly,
all suitable modifications and equivalents may be resorted to,
falling within the scope of the invention.
[0150] For example, in alternative embodiments of the invention,
the sample to be tested could include urine, serum, plasma, ocular
fluid or filtered whole blood. Suitable filtering systems for whole
blood could be incorporated into the cartridge. The screening
device could also used in other areas of immunodiagnostics. For
example, the screening device could be used to analyse the
concentration of tumour markers in the blood samples of patients
undergoing treatment for cancer. The screening device could also be
adapted for use to measure the levels of hormone, or therapeutic
drug present in a sample or to test for bacteria, viruses or other
microorganisms present in a variety of sample types. Alternatively
the screening device could be adapted to screen samples for
allergies.
[0151] It should be noted that the features described by reference
to particular figures and at different points of the description
may be used in combinations other than those particularly described
or shown. All such modifications are encompassed within the scope
of the invention as set forth in the following claims.
* * * * *