U.S. patent application number 10/461219 was filed with the patent office on 2003-11-13 for microdroplet dispensing methods for a medical diagnostic device.
Invention is credited to Harding, Ian A., Leung, Lewis, Renowitzky, Glen, Shartle, Robert Justice.
Application Number | 20030210287 10/461219 |
Document ID | / |
Family ID | 23803682 |
Filed Date | 2003-11-13 |
United States Patent
Application |
20030210287 |
Kind Code |
A1 |
Harding, Ian A. ; et
al. |
November 13, 2003 |
Microdroplet dispensing methods for a medical diagnostic device
Abstract
A medical diagnostic device has a non-absorbent substrate that
has a hydrophilic target area on which a reagent is deposited by
non-impact printing of microdroplets. During deposition, the device
is moved relative to the stream of microdroplets to form a
substantially uniform reagent layer on the substrate. The device is
particularly well adapted for measuring blood coagulation times. In
a preferred embodiment, coagulation times are determined by
monitoring the optical transmission of light through the target
area as an applied blood sample interacts with the reagent.
Inventors: |
Harding, Ian A.; (San Mateo,
CA) ; Shartle, Robert Justice; (Livermore, CA)
; Renowitzky, Glen; (San Lorenzo, CA) ; Leung,
Lewis; (Fremont, CA) |
Correspondence
Address: |
BOZICEVIC, FIELD & FRANCIS LLP
200 MIDDLEFIELD RD
SUITE 200
MENLO PARK
CA
94025
US
|
Family ID: |
23803682 |
Appl. No.: |
10/461219 |
Filed: |
June 13, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10461219 |
Jun 13, 2003 |
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09454196 |
Dec 3, 1999 |
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09454196 |
Dec 3, 1999 |
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09333765 |
Jun 15, 1999 |
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6521182 |
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60093421 |
Jul 20, 1998 |
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Current U.S.
Class: |
347/1 ;
427/2.13 |
Current CPC
Class: |
B01L 2200/0621 20130101;
B05D 3/00 20130101; G01N 33/5304 20130101; B01L 2300/0825 20130101;
G01N 33/558 20130101; G01N 2333/96447 20130101; G01N 33/54386
20130101; B01L 2300/0822 20130101; B01L 2400/0688 20130101; B01L
2300/0681 20130101; B01L 2300/087 20130101; B01L 2400/0406
20130101; B01L 3/5027 20130101; G01N 33/521 20130101; B01L
2300/0864 20130101; B01L 2300/0887 20130101; G01N 21/8483 20130101;
B01L 2200/16 20130101; G01N 33/525 20130101; G01N 33/86 20130101;
B01L 3/502707 20130101; G01N 33/4905 20130101; G01N 2333/7454
20130101; B01L 2400/0481 20130101; B01L 2200/12 20130101 |
Class at
Publication: |
347/1 ;
427/2.13 |
International
Class: |
B05D 003/00 |
Claims
We claim
1. A method for preparing a medical diagnostic reagent device,
comprising the steps of a) providing a non-absorbent substrate,
having on its surface at least one hydrophilic target area, b)
providing from a nonimpact print head onto a point within the
target area a pulsed stream of microdroplets of a diagnostic
reagent liquid. c) moving the stream relative to the substrate, and
d) repeating steps b) and c) at least enough times to provide a
substantially uniform layer of the liquid over the target area.
2. The method of claim 1, in which the substrate comprises a
substantially planar sheet.
3. The method of claim 1, in which the substrate comprises a
thermoplastic sheet.
4. The method of claim 1, in which each of the at least one target
areas has a water contact angle of no more than about
60.degree..
5. The method of claim 1, in which the print head is a thermal
ink-jet print head.
6. The method of claim 1, in which the reagent liquid comprises
thromboplastin.
7. The method of claim 2, in which the stream travels in a
direction that is substantially perpendicular to the substrate, and
the stream is moved relative to the substrate by moving the
substrate in a direction that is substantially perpendicular to the
direction of stream travel.
8. The method of claim 1, in which the stream passes through a hole
in a sheet that is positioned between the dispenser and
substrate.
9. The method of claim 8, in which the sheet has a hydrophobic
surface that faces the dispenser.
10. The method of claim 9, in which the reagent comprises a
colorant.
11. A diagnostic reagent device for measuring an analyte
concentration or characteristic of a biological fluid, including a
non-absorbent substrate comprising a) a sample application area for
accepting a sample of the biological fluid for analysis and b) a
predetermined hydrophilic reagent area, onto which has been
applied, by nonimpact printing, a diagnostic reagent liquid that
interacts with the sample to cause in the sample a
physically-measurable change that can be related to the analyte
concentration or characteristic of the fluid.
12. The device of claim 11, in which the sample application area
and reagent area substantially coincide.
13. The device of claim 11, further comprising means for conveying
the sample from the application area to the reagent area.
14. The device of claim 11, in which the sample application area is
hydrophilic.
15. The device of claim 11, in which the substrate comprises a
substantially transparent planar sheet.
16. The device of claim 11, in which the substrate comprises a
substantially transparent thermoplastic sheet.
17. The device of claim 11, in which the reagent liquid comprises
thromboplastin.
18. The device of claim 11, in which the reagent liquid comprises a
colorant.
19. The device of claim 13, in which the means for conveying the
sample from the application area to the reagent area comprises a
top layer, separated from the substrate by an intermediate layer
that has a through hole and adjoining channel cut into it, the top
layer, intermediate layer, and substrate forming a bladder that,
when compressed, and released causes in the channel a reduced
pressure that draws blood into the reagent area.
20. The device of claim 19, in which the top layer has a
hydrophobic surface facing the substrate, at least in the channel
and reagent area.
Description
CROSS-REFERENCE TO PRIOR APPLICATION
[0001] This application is a continuation-in-part of application
Ser. No. 09/333,765, filed Jun. 15, 1999.
BACKGROUND OF THE INVENTION
[0002] 1. Field of the Invention
[0003] This invention relates to a medical diagnostic device that
is prepared by nonimpact printing; more particularly, by nonimpact
printing of a reagent onto a hydrophilic surface of the device.
[0004] 2. Description of the Related Art
[0005] A variety of medical diagnostic procedures involve tests on
biological fluids, such as blood, urine, or saliva, and are based
on a change in a physical characteristic of such a fluid or an
element of the fluid, such as blood serum. The characteristic can
be an electrical, magnetic, fluidic, or optical property. When an
optical property is monitored, these procedures may make use of a
transparent or translucent device to contain the biological fluid
and a reagent. A change in light absorption of the fluid can be
related to an analyte concentration in, or property of, the fluid.
Typically, a light source is located adjacent to one surface of the
device and a detector is adjacent to the opposite surface. The
detector measures light transmitted through a fluid sample.
Alternatively, the light source and detector can be on the same
side of the device, in which case the detector measures light
scattered and/or reflected by the sample. Finally, a reflector may
be located at or adjacent to the opposite surface. A device of this
latter type, in which light is first transmitted through the sample
area, then reflected through a second time, is called a
"transflectance" device. References to "light" throughout this
specification and the appended claims should be understood to
include the infrared and ultraviolet spectra, as well as the
visible. References to "absorption" are meant to refer to the
reduction in intensity as a light beam passes through a medium;
thus, it encompasses both "true" absorption and scattering.
[0006] An example of a transparent test device is described in
Wells et al. W094/02850, published on Feb. 3, 1994. Their device
comprises a sealed housing, which is transparent or translucent,
impervious, and rigid or semi-rigid. An assay material is contained
within the housing, together with one or more assay reagents at
predetermined sites. The housing is opened and the sample
introduced just before conducting the assay. The combination of
assay reagents and analyte in the sample results in a change in
optical properties, such as color, of selected reagents at the end
of the assay. The results can be read visually or with an optical
instrument.
[0007] U.S. Pat. No. 3,620,676, issued on Nov. 16, 1971 to Davis,
discloses a calorimetric indicator for liquids. The indicator
includes a "half-bulb cavity", which is compressible. The bulb is
compressed and released to form a suction that draws fluid from a
source, through a half-tubular cavity that has an indicator
imprinted on its wall. The only controls on fluid flow into the
indicator are how much the bulb is compressed and how long the
indicator inlet is immersed in the source, while the bulb is
released.
[0008] U.S. Pat. No. 3,640,267, issued on Feb. 8, 1972 to Hurtig et
al., discloses a container for collecting samples of body fluid
that includes a chamber that has resilient, collapsible walls. The
walls are squeezed before the container inlet is placed into the
fluid being collected. When released, the walls are restored to
their uncollapsed condition, drawing fluid into and through the
inlet. As with the Davis device, discussed above, control of fluid
flow into the indicator is very limited.
[0009] U.S. Pat. No. 4,088,448, issued on May 9, 1978 to Lilja et
al., discloses a cuvette, which permits optical analysis of a
sample mixed with a reagent. The reagent is coated on the walls of
a cavity, which is then filled with a liquid sample. The sample
mixes with the reagent to cause an optically-detectable change.
[0010] The test devices described above and in the cited references
typically comprise a dry strip having a reagent coated on one or
more predetermined positions. Applying these reagents to their
intended positions on large numbers of these devices can, in
principle, be accomplished by standard printing processes; however,
nonimpact printing provides some distinct advantages. For example,
nonimpact printers can be smaller, lighter, and less expensive,
since they don't have to endure the repeated impact of print head
on substrate. They also permit the use of transparent substrates,
as required for optical devices that involve changes in light
transmission. Information on the varieties of nonimpact printing
appears in J. L. Johnson, Principles of Nonimpact Printing, 3d ed.,
Palatino Press, Irvine, Calif. 1998. (See, also, "No-splatter spray
makes better wafers," H. L. Berger, Machine Design, Feb. 5, 1998,
pp. 52-55). Among the varieties of nonimpact printing, ink-jet
printing has been identified as suitable for use in connection with
reagent fluids.
[0011] British Patent Specification, 1,526,708, published on Sep.
27, 1978, discloses a reagent test device that comprises a carrier
on which are printed two different substances, separated by a
"predetermined interspace." Ink-jet printing is one of the printing
techniques disclosed.
[0012] U.S. Pat. No. 4,877,745, issued on Oct. 31, 1989, to Hayes
et al., discloses a system for printing reagents onto a printing
medium by propelling droplets from a jetting tube and repeating the
process until a-desired configuration of the reagent is printed on
the medium. A piezo-electric print head was used.
[0013] U.S. Pat. No. 5,108,926, issued on Apr. 28, 1992, to Klebe,
discloses an apparatus for precisely locating cells on a substrate
by using an ink-jet printer either to deposit the cells directly
onto the substrate or to deposit cell adhesion materials. The
ink-jet printer used was a Hewlett-Packard Thinkjet.TM. printer,
which is a thermal ink-jet printer (see Hewlett-Packard Journal,
May, 1985).
[0014] U.S. Pat. No. 5,378,638, issued on Jan. 3, 1995, to Deeg et
al., discloses an analysis element for the determination of an
analyte in a liquid sample. The element is fabricated by ink-jet
printing of reagents in a series of "compartments," using a thermal
ink-jet print head.
[0015] Each of the references cited above are concerned, explicitly
or implicitly, with image spreading on the print medium, because
the sharpness of an image is degraded to the extent that the liquid
"ink" spreads across the surface before drying. For diagnostic
applications, sharp "images" are typically required, because
different reagents are positioned close together on a surface of a
device but must not come into contact (e.g., to react) until the
device is wetted by an applied sample.
SUMMARY OF THE INVENTION
[0016] The present invention provides a method for preparing a
medical diagnostic reagent device, comprising the steps of
[0017] a) providing a non-absorbent substrate, having on its
surface at least one hydrophilic target area,
[0018] b) providing from a nonimpact print head onto a point within
the target area a pulsed stream of microdroplets of a diagnostic
reagent liquid,
[0019] c) moving the stream relative to the substrate, and
[0020] d) repeating steps b) and c) at least enough times to
provide a substantially uniform layer of the liquid over the target
area.
[0021] A diagnostic reagent device of the present invention
measures an analyte concentration or characteristic of a biological
fluid and comprises
[0022] a) a sample application area for accepting a sample of the
biological fluid for analysis and
[0023] b) a predetermined hydrophilic reagent area, onto which has
been applied, by nonimpact printing, a diagnostic reagent liquid
that interacts with the sample to cause in the sample a
physically-measurable change that can be related to the analyte
concentration or characteristic of the fluid.
[0024] The sample application and reagent areas may coincide or,
alternatively, be spaced apart, with an intermediate path to convey
the sample. The measurement is generally, but not necessarily, made
when the sample is on the reagent area, and in the description
below, the measurement of interest is made when the sample is in
the reagent area.
[0025] The method is particularly well adapted for preparing a
device for measuring prothrombin time (PT time), with the target
area being coated with a reagent composition that catalyzes the
blood clotting cascade. Similarly, the diagnostic reagent strip of
the invention is particularly well adapted for measuring the PT
time of a whole blood sample.
[0026] As used in this specification and the appended claims, the
term "microdroplet" refers to droplets having a volume in the range
from about 1 picoliter to 1 microliter.
[0027] It is surprising that the hydrophilicity of the target area
provides superior results, since the hydrophilic surface would be
expected to spread the reagent that is deposited, which had been
thought to be undesirable.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] FIG. 1 is a plan view of a device of the present
invention.
[0029] FIG. 2 is an exploded view of the device of FIG. 1.
[0030] FIG. 3 is a perspective view of the device of FIG. 1.
[0031] FIG. 4 is a schematic of a meter for use with a device of
this invention.
[0032] FIG. 5 is a graph of data that is used to determine PT
time.
[0033] FIG. 6 is a plan view of an alternative embodiment of a
device of this invention.
[0034] FIG. 7 is a plan view of a coating prepared by the method of
the present invention.
[0035] FIG. 8 is a schematic of a nonimpact printing process of
this invention.
[0036] FIG. 9 is a graph that demonstrates an advantage of the
present invention.
DETAILED DESCRIPTION OF THE INVENTION
[0037] The medical diagnostic reagent device of this invention is
prepared by depositing a reagent upon a hydrophilic "reagent area"
of a non-absorbent substrate by a nonimpact printing process. The
device is of the type that relates a physical parameter of a
biological fluid, or an element of the fluid, to an analyte
concentration in the fluid or to a property of the fluid. Although
a variety of physical parameters--e.g., electrical, magnetic,
fluidic, or optical--can form the basis for the measurement, a
change in optical parameters is a preferred basis, and the details
that follow refer to an optical device. A preferred embodiment of
the device includes a planar substrate, such as a thermoplastic
sheet. The substrate has on its surface a sample application area
and the reagent area, in which the sample undergoes a change in an
optical parameter, such as light scattering. The substrate, or
"bottom layer," forms with "intermediate" and "top" layers a
bladder, to create a suction force to draw the sample into the
device, and a stop junction, to precisely stop flow after filling
the reagent area.
[0038] Preferably, the device is substantially transparent over the
reagent area, so that the area can be illuminated by a light source
on one side and the transmitted light measured on the opposite
side. The nonimpact-printed reagent causes the sample to undergo a
change, and the change in transmitted light is a measure of the
analyte or fluid property of interest. Alternatively, light that is
scattered from a fluid sample or light that passes through the
sample and is reflected back through a second time (by a reflector
on that opposite side) can be detected by a detector on the same
side as the light source.
[0039] This type of device is suitable for a variety of analytical
tests of biological fluids, such as determining biochemical or
hematological characteristics, or measuring the concentration in
such fluids of proteins, hormones, carbohydrates, lipids, drugs,
toxins, gases, electrolytes, etc. The procedures for performing
these tests have been described in the literature. Among the tests,
and where they are described, are the following:
[0040] (1) Chromogenic Factor XIIa Assay (and other clotting
factors as well): Rand, M. D. et al., Blood, 88, 3432 (1996).
[0041] (2) Factor X Assay: Bick, R. L. Disorders of Thrombosis and
Hemostasis: Clinical and Laboratory Practice. Chicago, ASCP Press,
1992.
[0042] (3) DRVVT (Dilute Russells Viper Venom Test): Exner, T. et
al., Blood Coag. Fibrinol., 1, 259 (1990).
[0043] (4) Immunonephelometric and Immunoturbidimetric Assays for
Proteins: Whicher, J. T., CRC Crit. Rev. Clin Lab Sci. 18:213
(1983).
[0044] (5) TPA Assay: Mann, K. G., et.al., Blood, 76, 755, (1990).;
and Hartshorn, J. N. et al., Blood, 78, 833 (1991).
[0045] (6) APTT (Activated Partial Thromboplastin Time Assay):
Proctor, R. R. and Rapaport, S. I. Amer. J. Clin. Path, 36, 212
(1961); Brandt, J. T. and Triplett, D. A. Amer. J. Clin. Path., 76,
530 (1981); and Kelsey, P. R. Thromb. Haemost. 52, 172 (1984).
[0046] (7) HbA1c Assay (Glycosylated Hemoglobin Assay): Nicol, D.
J. et al., Clin. Chem. 29, 1694 (1983).
[0047] (8) Total Hemoglobin: Schneck et al., Clinical Chem., 32/33,
526 (1986); and U.S. Pat. No. 4,088,448.
[0048] (9) Factor Xa: Vinazzer, H., Proc. Symp. Dtsch. Ges. Klin.
Chem., 203 (1977), ed. By Witt, I
[0049] (10) Colorimetric Assay for Nitric Oxide: Schmidt, H. H., et
al., Biochemica, 2, 22 (1995).
[0050] The present device is particularly well suited for measuring
blood-clotting time--"prothrombin time" or "PT time"--and details
regarding such a device appear below. The modifications needed to
adapt the device for applications such as those listed above
require no more than routine experimentation.
[0051] FIG. 1 is a plan view of a device 10 of the present
invention. FIG. 2 is an exploded view and FIG. 3 a perspective view
of the device. Sample is applied to sample port 12 after bladder 14
has been compressed. Clearly, the region of layer 26 and/or layer
28 that adjoins the cutout for bladder 14 must be resilient, to
permit bladder 14 to be compressed. Polyester of about 0.1 mm
thickness has suitable resilience and springiness. Preferably, top
layer 26 has a thickness of about 0.125 mm, bottom layer 28 about
0.100 mm. When the bladder is released, suction draws sample
through channel 16 to reagent area 18, which contains a
nonimpact-printed reagent 20. In order to ensure that reagent area
18 can be filled with sample, the volume of bladder 14 is
preferably at least about equal to the combined volume of channel
16 and reagent area 18. If reagent area 18 is to be illuminated
from below, layer 28 must be transparent where it adjoins reagent
area 18. For a PT test, reagent 20 contains thromboplastin that is
free of bulking reagents normally found in lyophilized
reagents.
[0052] As shown in FIGS. 1, 2, and 3, stop junction 22 adjoins
bladder 14 and reagent area 18; however, a continuation of channel
16 may be on either or both sides of stop junction 22, separating
the stop junction from reagent area 18 and/or bladder 14. When the
sample reaches stop junction 22, sample flow stops. For PT
measurements, it is important to stop the flow of sample as it
reaches that point to permit reproducible "rouleaux formation"--the
stacking of red blood cells--which is an important step in
monitoring blood clotting using the present invention. The
principle of operation of stop junctions is described in U.S. Pat.
No. 5,230,866, incorporated herein by reference.
[0053] As shown in FIG. 2, all the above elements are formed by
cutouts in intermediate layer 24, sandwiched between top layer 26
and bottom layer 28. Preferably, layer 24 is double-sided adhesive
tape. Stop junction 22 is formed by an additional cutout in layer
26 and/or 28, aligned with the cutout in layer 24 and sealed with
sealing layer 30 and/or 32. Preferably, as shown, the stop junction
comprises cutouts in both layers 26 and 28, with sealing layers 30
and 32. Each cutout for stop junction 22 is at least as wide as
channel 16. Also shown in FIG. 2 is an optional filter 12A to cover
sample port 12. The filter may separate out red blood cells from a
whole blood sample and/or may contain a reagent to interact with
the blood to provide additional information. A suitable filter
comprises an anisotropic membrane, preferably a polysulfone
membrane of the type available from Spectral Diagnostics, Inc.,
Toronto, Canada. Optional reflector 18A may be on, or adjacent to,
a surface of layer 26 and positioned over reagent area 18. If the
reflector is present, the device becomes a transflectance
device.
[0054] The method of using the strip of FIGS. 1, 2, and 3 can be
understood with reference to a schematic of the elements of a meter
shown in FIG. 4, which contemplates an automated meter.
Alternatively, manual operation is also possible. (In that case,
bladder 14 is manually depressed before sample is applied to sample
port 12, then released.)
[0055] The first step the user performs is to turn on the meter,
thereby energizing strip detector 40, sample detector 42,
measurement system 44, and optional heater 46. The second step is
to insert the strip. Preferably, the strip is not transparent over
at least a part of its area, so that an inserted strip will block
the illumination by LED 40a of detector 40b. (More preferably, the
intermediate layer is formed of a non-transparent material, so that
background light does not enter measurement system 44.) Detector
40b thereby senses that a strip has been inserted and triggers
bladder actuator 48 to compress bladder 14. A meter display 50 then
directs the user to apply a sample to sample port 12 as the third
and last step the user must perform to initiate the measurement
sequence.
[0056] The empty sample port is reflective. When a sample is
introduced into the sample port, it absorbs light from LED 42a and
thereby reduces the light that is reflected to detector 42b. That
reduction in light, in turn, signals actuator 48 to release bladder
14. The resultant suction in channel 16 draws sample through
reagent area 18 to stop junction 22. Light from LED 44a passes
through reagent area 18, and detector 44b monitors the light
transmitted through the sample as it is clotting. When there are
multiple reagent areas, measurement system 44 includes an
LED/detector pair (like 44a and 44b) for each reagent area.
Analysis of the transmitted light as a function of time (as
described below) permits a calculation of the PT time, which is
displayed on the meter display 50. Preferably, sample temperature
is maintained at about 37.degree. C. by heater 46.
[0057] FIG. 5 depicts a typical "clot signature" curve in which the
current from detector 44b is plotted as a function of time. Blood
is first detected in the reagent area by 44b at time 1. In the time
interval A, between points 1 and 2, the blood fills the reagent
area. The reduction in current during that time interval is due to
light scattered by red cells and is thus an approximate measure of
the hematocrit. At point 2, sample has filled the reagent area and
is at rest, its movement having been stopped by the stop junction.
The red cells begin to stack up like coins (rouleaux formation).
The rouleaux effect allows increasing light transmission through
the sample (and less scattering) in the time interval between
points 2 and 3. At point 3, clot formation ends rouleaux formation
and transmission through the sample reaches a maximum. The PT time
can be calculated from the interval B between points 1 and 3 or
between 2 and 3. Thereafter, blood changes state from liquid to a
semi-solid gel, with a corresponding reduction in light
transmission. The reduction in current C between the maximum 3 and
endpoint 4 correlates with fibrinogen in the sample.
[0058] FIG. 6 depicts a preferred embodiment of the present device.
It is a multi-channel device that includes a bypass channel 52.
Bypass channel 52 provides a path for sample to travel after sample
has been drawn into reagent areas 118, 218, and 318. Sample is
drawn into the bypass channel by the reduced pressure on the
bladder side of stop junction 122. Sample flow stops when the
ambient pressure is equalized on both sides of the stop junction.
Reagent area 118 contains thromboplastin. Preferably, reagent areas
218 and 318 contain controls, more preferably, the controls
described below. Area 218 contains thromboplastin, bovine eluate,
and recombinant Factor VIIa. The composition is selected to
normalize the clotting time of a blood sample by counteracting the
effect of an anticoagulant, such as warfarin. Reagent area 318
contains thromboplastin and bovine eluate alone, to partially
overcome the effect of an anticoagulant. Thus, three measurements
are made on the strip. PT time of the sample, the measurement of
primary interest, is measured on area 118. However, that
measurement is validated only when measurements on areas 218 and
318 yield results within a predetermined range. If either or both
of these control measurements are outside the range, then a retest
is indicated. Extended stop junction 122 stops flow in all three
reagent areas.
[0059] The device pictured in FIGS. 1 and 2 and described above is
preferably formed by laminating thermoplastic sheets 26 and 28 to a
thermoplastic intermediate layer 24 that has adhesive on both of
its surfaces. The cutouts that form the elements shown in FIG. 1
may be formed, for example, by laser- or die-cutting of layers 24,
26, and 28.
[0060] The reagent area 18 on bottom layer 28 is defined by the
cutout in intermediate layer 24. Preferably; the bottom surface of
top layer 26, facing bottom layer 28, is hydrophobic, at least in
the region of channel 16 and reagent area 18. The surface of
reagent area 18 is hydrophilic. Preferably, the surface of sample
port 12 is hydrophilic as well, to facilitate filling of the
device; i.e., moving the sample from port 12 to reagent area 18. A
convenient way to have hydrophilic sample and reagent areas is to
have the entire surface of bottom layer 28 be hydrophilic.
Commercially available thermoplastic films having suitably
hydrophilic surfaces include 3M 9962 Antifog Film ("Antifog"),
available from Medical Specialties, 3M Health Care, St. Paul,
Minn.; FMC GelBond Film, available from Bio Whittaker Molecular
Applications, Rockland, Me.; polyethylene terephthalate (PET) film,
whose surface has been flame-corona- or plasma-treated; ionomer
film; and other conventional thermoplastic films having hydrophilic
surfaces or coatings. The Antifog is PET film coated with a
3M-proprietary coating and is the preferred substrate material.
[0061] In determining the suitability of a substrate for the
present device and method, the surface hydrophilicity can be
determined in several different ways.
[0062] Contact angle is nominally the angle between the edge of a
drop of fluid (usually purified water) that sits atop a wettable
surface and the surface itself. The method for measuring the
contact angle has been standardized, and can be carried out using
manual or automated equipment. (ASTM Test Method D5946-96, Standard
Test Method for Corona-Tested Polymer Films Using Water Contact
Angle Measurements.) The data can generally by considered accurate
and reproducible when the measured angle is greater than
25.degree., and films are considered quite wettable if the contact
angle is about 60.degree. or less. The angles measured for Antifog
were about 25.degree..
[0063] Wetting tension is measured by spreading solutions of known
surface tension onto a surface to be tested and observing if the
solutions "bead up." (ASTM Test Method D2578-94, Standard Test
Method for Wetting Tension of Polyethylene and Polypropylene
Films). Beading up indicates that internal liquid attractive forces
overcome adsorptive attraction of the surface. The solutions are
calibrated in units of dynes/cm, and are referred to as dyne
solutions. They are commercially available in the range of 30 to 60
dynes/cm. A surface is tested starting with the lowest value
solution and progressing to the highest. A surface is assigned the
dyne/cm value corresponding to that solution that remains spread
out for approximately two seconds. Since Antifog wetted out all the
solutions, it has been characterized as having a surface wetting
tension greater than 60 dynes/cm.
[0064] 3M's Medical Specialties Department has developed a wetting
test to characterize water-wetting of film. (3M SMD #6122, Wetting
Test, Dec. 4, 1998--available from 3M Center, St. Paul, Minn.
55144-1000.) The test involves careful placement of an aqueous dye
solution onto a surface, drying it, and measuring the diameter of
the dried spots. The data collected were generally in the 35 to 40
point range, which indicates a very wettable surface.
[0065] Based on the measurements described above, we conclude that
the Antifog surface is extremely hydrophilic. When a surface is
adequately hydrophilic, then reagent droplets spread over the
surface and, providing sufficient droplets are deposited, form a
substantially uniform layer of the reagent over the desired area.
As used in this specification and the appended claims, the term
"substantially uniform" should not be construed as necessarily
suggesting that the surface coating thickness is the same over the
entire target area, nor even that the entire surface is coated.
[0066] FIG. 7 depicts a plan view of part of a typical coated
target area. Note that part of the surface (A) remains uncoated,
although most of the surface (B) is coated. Preferably, at least
about 80% of the target area is coated. Preferably, thickness
variations in the coated areas (B) are minimized; e.g., thickest
region less than three times the average thickness of the coated
area. Average coating thickness in coated areas is generally about
0.1 micrometer-about 1 micrometer, depending on the nature of the
reagent and the particular application.
[0067] FIG. 8 depicts a schematic of an apparatus for nonimpact
printing of reagent onto the reagent area of a substrate of the
present invention. Print head 60 repeatedly ejects a stream of
reagent droplets onto web 62, which moves in the direction shown by
the arrow. Optional masks 64 and 66 ensure that the droplet stream
only reaches web 62 in reagent areas 18.
[0068] To control the printing, mask 66; i.e., the mask closest to
print head 60, optionally has a hydrophobic surface 68 facing the
print head. Reagent from the multiple dispenser nozzles of print
head 60 forms multiple reagent dots on mask surface 68. Because the
surface is hydrophobic, the dots remain isolated and can be
individually viewed by a downstream optical system 70. The
hydrophilicity of surface 18 causes the droplets arriving on that
surface to spread and/or coalesce, so it is more difficult for
optical system 70 to detect individual dots directly on the reagent
area.
[0069] Optical system 70 can detect and, if desired, reject
defective product. For example, an absence of dots may indicate
that one or more dispenser nozzles are defective. Among the
suitable optical detection methods are dark field microscopy,
shadowing, patterning, laser illumination, etc. Optionally, a
colorant, or a fluorescent dye, can be added to the reagent to make
it more easily visible to optical system 70. For example, methylene
blue dye, added to a reagent to about 0.1% final concentration,
makes the reagent visible to an optical system, without
substantially altering the measurements made with the reagent.
[0070] Print head 60 may be any nonimpact print head known in the
art, including ultrasonic, electrographic, ion projection, etc.
Preferably, print head 60 is an ink-jet print head, more
preferably, a thermal ink-jet print head.
[0071] The following examples demonstrate the present invention in
its various embodiments, but are not intended to be in any way
limiting.
EXAMPLE 1 (COMPARATIVE EXAMPLE)
[0072] Two strips of the type described above for PT measurements
were prepared (see FIGS. 1-3). The difference between the strips
was that strip A had a bottom layer 28 of untreated polyethylene
terephthalate;, while strip B had a bottom layer 28 of FMC GelBond
Film. A blood sample was applied to each strip and PT measurements
made in an apparatus of the type depicted in FIG. 4. FIG. 9 depicts
the resultant clotting curves. The curve for strip A has a
relatively flat peak (corresponding to peak 3 in FIG. 5). The
flatness of the peak limits the precision of the resultant PT
calculation. By contrast, the curve for strip B has a much sharper
peak, which permits much greater precision. (Note that the PT times
for the samples measured with the two strips are different.)
EXAMPLE 2
[0073] A device of this invention is made by first passing a
double-sided adhesive tape (RX 675SLT, available from Scapa Tapes,
Windsor, CT) sandwiched between two release liners into a
laminating and rotary die-cutting converting system. The pattern
shown in FIG. 2, with the exception of the stop junction, is cut
through the top release liner and tape, but not through the bottom
release liner, which is then removed as waste, along with the
cutouts from the tape. 3M Antifog Film is laminated to the exposed
bottom side of the tape. Reagent (thromboplastin, available from
Ortho Clinical Diagnostics, Raritan, N.J.) is then printed onto the
reagent area (18) of the film by thermal ink-jet printing, using
printing heads 51612A from Hewlett Packard, Corvallis, Oreg. A
sample port is cut in untreated polyester film (AR1235, available
from Adhesives Research, Glen Rock, Pa.) and then laminated, in
register, to the top of the double-sided tape (after removing the
release layer). A die then cuts the stop junction through the three
layers of the sandwich. Finally, strips of single-sided adhesive
tape--Catalog No. 9843 (MSX4841), available from 3M, St. Paul,
Minn.--are applied to the outside of the polyester layers to seal
the stop junction.
EXAMPLE 3
[0074] A procedure that is similar to the one described in Example
1 is followed to make a strip of the type depicted in FIG. 6.
Reagent that is thermal ink-jet printed onto areas 118P, 218P, and
318P is, respectively, thromboplastin; thromboplastin, bovine
eluate, and recombinant Factor VIIa; and thromboplastin and bovine
eluate alone. The bovine eluate (plasma barium citrate bovine
eluate) is available from Haemotologic Technologies, Burlington,
Vt.; and recombinant Factor VIIa from American Diagnostica,
Greenwich, Conn.
[0075] Measurements made on a whole blood sample using the strip of
this Example yield a curve of the type shown in FIG. 5 for each of
the reagent areas. The data from the curves for the controls
(reagent areas 218P and 318P) are used to qualify the data from the
curve for reagent area 118P. As a result, the PT time can be
determined more reliably than can be done with a strip having a
single reagent area.
* * * * *