U.S. patent application number 10/968524 was filed with the patent office on 2007-03-08 for patient sample classification based upon low angle light scattering.
This patent application is currently assigned to Ortho-Clinical Diagnostics, Inc.. Invention is credited to Zhong Ding, Merrit N. Jacobs, Michael W. Sundberg.
Application Number | 20070054405 10/968524 |
Document ID | / |
Family ID | 34396608 |
Filed Date | 2007-03-08 |
United States Patent
Application |
20070054405 |
Kind Code |
A1 |
Jacobs; Merrit N. ; et
al. |
March 8, 2007 |
Patient sample classification based upon low angle light
scattering
Abstract
An apparatus for classifying a liquid patient sample includes at
least one sample container having a quantity of a sample that is
aggressively acted upon so as to create a flow field. A measurement
mechanism includes at least one low angle light emitter aligned
with a measurement window of the at least one sample container and
a detector oppositely disposed relative to the measurement window.
Measurement of the scattered light detects particle characteristics
of a moving flow field from the sample to determine, for example,
the amount of agglutination of the sample so as to perform blood
typing or other classifications without spatial separation.
Inventors: |
Jacobs; Merrit N.;
(Fairport, NY) ; Sundberg; Michael W.; (Pittsford,
NY) ; Ding; Zhong; (Pittsford, NY) |
Correspondence
Address: |
WALL MARJAMA & BILINSKI
250 SOUTH CLINTON STREET
SUITE 300
SYRACUSE
NY
13202
US
|
Assignee: |
Ortho-Clinical Diagnostics,
Inc.
Rochester
NY
|
Family ID: |
34396608 |
Appl. No.: |
10/968524 |
Filed: |
October 19, 2004 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60513753 |
Oct 23, 2003 |
|
|
|
Current U.S.
Class: |
436/69 |
Current CPC
Class: |
G01N 15/042 20130101;
G01N 15/0211 20130101; G01N 33/80 20130101; G01N 33/4905 20130101;
G01N 15/05 20130101 |
Class at
Publication: |
436/069 |
International
Class: |
G01N 33/86 20060101
G01N033/86 |
Claims
1. A method of classifying an agglutination reaction, said method
comprising the steps of: providing a mixture of a blood sample and
an agglutination reagent into the confines of a sample container;
moving the mixture within the sample container such that an
agglutination reaction occurs and that the agglutinated cells are
moved as a flow field into a measurement window of the sample
container; aiming a light source into the sample container and
probing the flow field with the light beam; and detecting the
number and size of particles in the light to determine the amount
of agglutination of said sample.
2. A method as recited in claim 1, wherein the moving step is
performed by centrifugation.
3. A method as recited in claim 1, wherein said moving step is
performed by vertical movement of the sample within a reaction
vessel serving as said sample container.
4. A method as recited in claim 3, wherein said reaction vessel
includes at least one transition zone having a smaller inside
diameter than an adjacent portion through which the sample is
moved.
5. A method as recited in claim 1, wherein said light beam is
scattered by particles contained in said moving flow field, said
scattered light being detected in said detecting step.
6. A method as recited in claim 2, wherein said sample container
contains at least one fluid having a specific viscosity and
specific gravity to effectively exclude the sample prior to
centrifugation thereof.
7. A method as recited in claim 1, wherein said aiming and
detecting steps are performed using a particle measurement
system.
8. A method of classifying a patient sample, said method comprising
the steps of: providing a patient sample in a sample container;
moving a quantity of sample fluid contained within said sample
container so as to create a moving flow field; aiming a light
source into the sample container and probing the flow field with
the light beam; and detecting particle characteristics of said flow
field in order to effectively classify said sample.
9. A method as recited in claim 8, wherein said patient sample is a
blood sample.
10. A method as recited in claim 9, including the step of providing
a mixture of a blood sample and at least one agglutination reagent
into the confines of said sample container prior to said mixing
step wherein said moving step includes moving the mixture of said
blood sample and said at least one agglutination reagent in order
to create an agglutination reaction.
11. A method as recited in claim 10, wherein said moving step is
performed using centrifugation.
12. A method as recited in claim 10, wherein said moving step is
performed using agitation of said sample container.
13. A method as recited in claim 12, wherein said sample container
includes at least one chamber having a smaller inside diameter than
an adjacent chamber of said container through which the mixture is
moved.
14. A method as recited in claim 11, including the step of
providing at least one fluid having a specific viscosity and
specific gravity to effectively exclude the sample prior to
centrifugation thereof.
15. A method as recited in claim 11, wherein said particle
characteristics include the size distribution of agglutinated
particles in said flow field.
16. A method as recited in claim 9, wherein said method includes
the step of blood typing based on the particle characteristics of
said sample.
17. A method as recited in claim 9, wherein said detecting step
includes the steps of detecting antigens from said sample.
18. An apparatus for classifying a patient sample, said apparatus
comprising: at least one sample container; means for moving a
quantity of sample fluid contained within said at least one sample
container so as to create a moving flow field; and a measurement
mechanism including a light emitter aligned with a measurement
window of said at least one sample container and a light detector
oppositely disposed relative to said measurement window for
detecting scattered light from an emitted light beam, wherein said
detector detects particle characteristics of said flow field in
order to effectively classify said sample.
19. An apparatus as recited in claim 18, wherein said moving means
includes a centrifuge.
20. An apparatus as recited in claim 18, wherein said moving means
includes means for agitating said at least one sample
container.
21. An apparatus as recited in claim 18, wherein said light
detector is aligned relative to the axis of said light beam of said
light source by an angle that is less than about 10 degrees.
22. An apparatus as recited in claim 20, wherein said sample
container includes at least one chamber having an inside diameter
which is smaller than an adjacent chamber to promote mixing.
23. An apparatus as recited in claim 18, wherein said moving means
and said measurement system are not integrally provided on said
apparatus, wherein said measurement system can be used to detect
scattered light after said at least one sample container has been
removed from said moving means.
24. An apparatus as recited in claim 18, wherein said moving means
and said measurement system are integrally provided on said
apparatus such that measurements can be timely made following said
mixing step.
25. An apparatus as recited in claim 18, wherein said
classification includes detection of the amount of agglutination of
a patient sample.
26. An apparatus as recited in claim 18, wherein said
classification includes detection of the amount of at least one
antigen in a patient sample.
27. An apparatus as recited in claim 25, wherein said
classification further includes blood typing.
Description
CROSS REFERENCE TO RELATED APPLICATION
[0001] This application is based upon a provisional application,
U.S. Ser. No. 60/513,753 filed Oct. 23, 2003, the entire contents
of which are incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The invention is directed to the field of clinical chemistry
and more particularly to apparatus and a related method for blood
typing patient samples.
BACKGROUND OF THE INVENTION
[0003] Currently known column blood-typing systems, including, for
example, the AutoVue systems manufactured by Ortho Clinical
Diagnostics Inc., of Raritan, N.J., among others, were originally
optimized for manual testing. As a consequence, a number of
characteristics of the systems, while supporting the manual
determination of a positive reaction, do not support facile
automation.
[0004] First, agglutination strength is determined in a manner that
is optimized for a subjective visual read. This determination is
based upon a perceived distribution of red color from red blood
cells, RBCs, in an elongated column, with an agglutinated sample
being characterized by red color that is distributed above the
bottom of the column, with a non-agglutinated sample having red
color localized at the bottom thereof.
[0005] Automated reading of the distribution of red color is
complex and requires a vision system that is coupled with
relatively sophisticated software algorithms in order to
appropriately classify the wide range of color distribution
patterns that can be encountered. Such a technique is described,
for example, in U.S. Pat. Nos. 5,594,808 and 5,768,407.
[0006] In these systems, a centrifugation step of relatively long
duration (e.g., approximately 15 to 20 minutes) is typically
required in order to achieve a necessary spatial separation of the
agglutinated from the unagglutinated cells, including both
incubation and centrifugation times.
[0007] Analysis of agglutination patterns however, is often an
urgent procedure, for example in the case of injury, and the
elimination of time in the blood typing process is therefore highly
significant and greatly desired in the field.
[0008] A technique that permits agglutination to be detected
without centrifugation has been more recently described by
Applicants' in U.S. Publication Nos. US 2002/0076826 and US
2002/0081747, each of which is incorporated by reference in its
entirety. This technique defines apparatus and a method for
aspirating within a probe tip and is defined such that most of the
liquid is forced to move past a defined transition zone between two
different inside diameters of the tip to cause rotational mixing of
the liquid. This method is useful in providing agglutination of
blood, which can then in turn be used in blood typing. The
classification and determination of agglutination strength,
however, regardless of the technique used to cause separation, is
the same as described above, e.g., thereby requiring spatial
separation of the agglutinated/unagglutinated cells.
SUMMARY OF THE INVENTION
[0009] It is therefore a primary object of the present invention to
alleviate or substantially minimize the above-noted deficiencies of
the prior art.
[0010] It is another primary object of the present invention to
provide a blood typing methodology that is more efficient in terms
of time and accuracy than any previously known typing
technique.
[0011] It is yet another primary object of the present invention to
provide an agglutination reaction detection system and a related
method that is not solely dependent upon spatial separation in
order to be able to detect the amount of agglutination of a sample
liquid.
[0012] Therefore and according to a preferred aspect of the present
invention, there is described a method of classifying a patient
sample, said method including the steps of:
[0013] providing a mixture of a blood sample and an agglutination
reagent into the confines of a sample container;
[0014] moving the mixture within the sample container such that an
agglutination reaction occurs and that the agglutinated cells are
moved as a flow field past a measurement window of the sample
container;
[0015] aiming a light source into the sample container and probing
the flow field with the light beam; and
[0016] detecting the number and size of particles in the light beam
to determine the amount of agglutination of said sample.
[0017] According to one technique, the mixture is moved by means of
a vertical (e.g., up and down) movement of the sample within a
reaction vessel serving as the sample container in order to mix the
sample and to move the agglutinated cells through at least one
defined transition zone of the vessel, the transition zone having a
smaller inside diameter than the adjacent portion through which the
sample is moved under the force of gravity. In this manner,
agglutinated material is separated from non-agglutinated
material.
[0018] According to another technique, the sample container can be
centrifuged using at least one fluid having a viscosity and
specific gravity that effectively excludes the sample prior to
centrifugation, but which allows the particles to enter when the
centrifugal force is applied, while excluding small molecules and
controlling the rate at which the material moves within the fluid
under the centrifugal field.
[0019] According to the present technique, a cloud of cells are
formed in the sample container. A low-angle particle measurement
system comprising at least one light emitter and at least one light
detector is used to detect the agglutination in the cloud of cells
at specified times after initiating the reaction.
[0020] Preferably, the sample container is elongated in
configuration and includes a flat planar wall defining the
measurement window for permitting the light beam to be effectively
scattered by the particles contained in the moving flow field.
According to another preferred aspect of the invention, there is
described an apparatus for classifying a patient sample, said
apparatus including:
[0021] at least one sample container;
[0022] means for moving a quantity of sample fluid contained within
said at least one sample container so as to create a moving flow
field; and
[0023] a measurement mechanism including a light emitter aligned
with a measurement window of said at least one sample container and
a light detector oppositely disposed relative to said measurement
window for detecting scattered light from an emitted light beam,
wherein said detector detects particle characteristics of said flow
field in order to effectively classify said sample.
[0024] Preferably, the light source emits a beam that scatters at
low angles based upon the number and size of particles in a
scanning measurement volume wherein the aligned detector receives
the scattered light and through processing logic contained therein
is able to detect the amount of agglutination based on the detected
particle distribution, so as to perform blood typing or other
detection analyses of a patient sample.
[0025] An advantage of the herein described measurement system and
method is that blood typing and other detection analyses can be
performed in a much more time effective manner than previously
known techniques. Moreover, the newly described technique does not
require the use of an inert bead matrix or other similar means as
typically required, for example, in agglutination detection
processes that require spatial separation of agglutinated and
unagglutinated cells, such as those described, for example, in U.S.
Pat. Nos. 5,512,432 and 6,203,706B1, among others that require
spatial separation in order to effectuate a visual determination of
agglutination strength.
[0026] It will be readily apparent that the above method can be
used to detect other target antibodies or antigens, proteins,
viruses, or bacteria in a similar manner by producing a cloud of
particles in a reaction and determining the degree of agglutination
using a low angle light scattering means and a suitable measurement
system to determine the particle distribution from a scanned beam
impinging upon the moving flow field.
[0027] An advantage provided by the apparatus and method of the
present invention is that blood typing and other forms of
classification by reaction can be performed in an extremely fast
and efficient manner without requiring spatial separation.
[0028] Yet another advantage of the present invention is that the
herein described system and method can be added to already existing
equipment without significant modification thereto.
[0029] Yet another advantage of the present invention is that the
above described method provides an effective qualitative
determination of agglutination strength.
[0030] These and other objects, features and advantages will be
apparent from the following Detailed Description which should be
read in conjunction with the accompanying drawings.
BRIEF DESCRIPTION OF THE DRAWINGS
[0031] FIG. 1 is a cross-sectional view of a prior art test tube
cartridge containing multiple samples showing the patterns
occurring in positive, weakly positive, and negative agglutination
reactions for column agglutination assays;
[0032] FIG. 2 is a diagrammatical view illustrating the effects of
light scattering in a liquid medium from an incident beam;
[0033] FIG. 3 depicts a diagrammatic view of a low angle light
scattering/particle detection system for use in the present
invention, the system being depicted to shown unscattered light
through a liquid medium;
[0034] FIG. 4 depicts a partial, diagrammatic view of the low angle
light scattering/particle detection system of FIG. 3, as taken
during scattering events such as those previously depicted in FIG.
2;
[0035] FIG. 5 is a perspective view of a measurement system in
accordance with a first embodiment of the present invention;
[0036] FIG. 6 is a diagrammatic view of a measurement system in
accordance with a second embodiment of the present invention;
[0037] FIG. 7 is a side elevational view, taken is section of a
sample container made in accordance with a preferred embodiment of
the present invention;
[0038] FIG. 8 is the side elevational view of the sample container
of FIG. 7, depicting the movement of a flow field in relation to
the measurement system of FIG. 3;
[0039] FIG. 9 is a side elevational view of a sample container made
in accordance with the present invention;
[0040] FIG. 10 is a side elevational view of the sample container
of FIG. 9; and
[0041] FIG. 11 is a flow chart illustrating a method for performing
antibody detection in accordance with another preferred embodiment
of the invention using the measurement system of FIG. 5.
DETAILED DESCRIPTION
[0042] The following description relates to certain preferred
embodiments of the present invention, and to a particular
methodology for blood typing by detection of agglutination strength
by means of a low angle light scattering/detection system. As will
be readily apparent from the discussion, the inventive concepts
described herein can also be suitably applied to other reaction
processes in addition to blood typing to detect antigens,
antibodies, proteins, viruses, and the like wherein a reaction can
create a moving flow field. In addition, such terms as "top",
"bottom", "lateral", "above", "below" and the like are also used in
order to provide a convenient frame of reference for use with the
accompanying drawings. These terms, unless stated specifically
otherwise, however, are not intended to be limiting of the present
invention.
[0043] Referring to FIG. 1, the results of a prior art blood typing
system are herein described briefly for purposes of background. In
this system, as more completely described in U.S. Pat. No.
5,552,064 incorporated entirely by reference herein, a series of
tubular sample containers 14, 16, 18, 20, 22 are disposed within a
cartridge 10. Each of the tubular sample containers includes a
suspended matrix 30 of substantially noncompressible inert
microparticles that permit movement of nonagglutinated reactants,
such as red blood cells, while constraining agglutinated reactants.
The matrix 30 is typically suspended within a gel in a lower
portion 25 of the container, the gel having a density that is
slightly lower than of the red blood cells to promote movement
therethrough. Typically and for ABO blood typing, a sample and
agglutination reactant is first received within an upper portion 23
or chamber of each container in the form of serum and cells that
are incubated prior to movement through the matrix 30 of suspended
particles. The upper portion 23 is separated from the lower portion
25 by means of a separating orifice having a diameter permitting
the passage of cells therethrough, the serum and cell reagent being
applied so as to create an air pocket or bubble between the upper
portion and the reminder of the container. A user then applies a
sufficient force, typically centrifugation, to effect this movement
wherein a band of agglutinated reactants is formed above the matrix
30 for visual or automated detection. The container may also have
an initial reaction zone 21.
[0044] As noted, FIG. 1 illustrates a range of spatial separation
patterns that are representative of positive, weakly positive, and
negative agglutinations. Tubular sample container 12 demonstrates a
strong positive reaction with a firm band 19 of agglutinates.
Container 14 shows a positive reaction that is somewhat weaker than
that shown in container 12, as the agglutinate band has broken
apart into smaller agglutinates. Container 16 demonstrates a weaker
positive reaction with a smaller quantity of agglutinates being
distributed throughout the middle portion of the matrix, and even
settling on the bottom, as in container 18. Container 20 depicts a
very weak positive reaction, with most of the cells being collected
on the bottom. Finally, tubular sample container 22 depicts a clear
negative reaction, with a button of cells 26 located on the bottom
of the container, and no agglutinates dispersed within the matrix
30.
[0045] The present invention relates to use of a technique of
low-angle light scattering to provide a more accurate and
repeatable determination of agglutination without requiring spatial
separation. The basic theory of this technique follows:
[0046] Low-angle light scattering is generally based on the
principle that the absorption coefficient and the volume scattering
function (VSF) completely characterize how a beam of laser light
will propagate through water or other fluid. In brief, this
technique measures the intensity of light that is scattered through
a range of small angles from the original direction of propagation
as a result of particles in a detection area. Typically, the pick
up angle of the incident (e.g., the scattering angle) varies
between 0.1 to about 10 degrees.
[0047] In theory, the properties of an incident light beam into a
liquid medium, such as water, is illustrated in FIG. 2, wherein a
narrow, monochromatic collimated beam of light of power .PHI..sub.i
becomes incident upon a volume of liquid, having a thickness
designated herein as .DELTA.r. Some part of the incident power
.PHI..sub.s, (.psi.) of the beam is scattered into a solid angle
.DELTA..OMEGA. that is centered upon the scattering angle .psi..
The resulting function, referred to as the Volume Scattering
Function (VSF) is then defined by the following relation: VSF
.function. ( .psi. ) = .PHI. s .function. ( .psi. ) .PHI. i .times.
.DELTA. .times. .times. r .times. .times. .DELTA..OMEGA.
##EQU1##
[0048] Referring to FIGS. 3 and 4, a low angle scattering/particle
measurement system is shown for use in the present invention. A
monochromatic light beam 33 is formed by collimating the output of
a diode or other form of laser or other type of light emitter 32.
Scattered light is detected in this particular instance by means of
a detector 36 that is defined by a predetermined number of
coaxially arranged rings 38 allowing the passage of unscattered
light. FIG. 3 illustrates an unscattered light beam 33 that is
initially collimated by means of a lens 33 and is focused onto the
center of the detector 36 through a corresponding lens 37. As shown
more clearly in FIG. 4, each detector ring 38 is disposed so as to
collect the portion of the light beam that is scattered into a
particular solid angle .DELTA..OMEGA. and defined by a narrow range
of scattering angles (.psi.) relative to the medium, typically on
the order of about 0.1 to about 20 degrees. The herein described
system is from the LISST Series Particle Measurement Systems,
manufactured by Sequoia Scientific, Inc. of Redmond, Wash.
[0049] According to the invention, measurement of the Volume
Scattering Function (VSF) over the above noted range of pick up
angles can therefore be used, by mathematical processing to obtain
the size distribution and the concentration of suspended particles
in the liquid using a processor 41. The above system can detect
particles as small as those having a size in the single micron
range and therefore can detect the quantity of agglutinated and
nonagglutinated materials in an assay.
[0050] Because of its optical nature, the above-described technique
is facilitated if the entrance and exit surfaces of a sample
container are preferably approximately planar, made from a light
transmissive material, and are substantially perpendicular to the
angle of incident illumination in order to reduce artifacts that
may result from scatter at the container surfaces.
[0051] Referring to FIGS. 5 and 6, two different embodiments are
herein described for facilitating agglutination and localizing the
particulate materials preferably in conjunction with the low angle
scattering/particle detection system of FIGS. 3 and 4. It should be
readily discernible, however, that other techniques can be employed
within the inventive scope of the present invention.
[0052] Referring first to FIG. 5, a low angle light
scattering/particle measuring system having a laser emitter 32 and
a detector 36, shown only schematically, but as described with
regard to FIGS. 3 and 4, is aligned relative to a sample container
43 and more specifically to a measurement window 45 of the
container. The lenses 35, 37, FIG. 3, of the measuring system are
not shown in this view for reasons of clarity.
[0053] The sample container 43 for purposes of this embodiment is
defined by three adjacent chambers 51, 53, 55 each linked by a
common wall, preferably, each of the chambers being defined by a
different inside diameter D1, D2 and D3, respectively. As shown,
each of D.sub.2 and D.sub.3 are substantially larger than D.sub.1.
In order to promote mixing, transition zones 44, 48 are created
between adjacent chambers so as to promote an agglutination
reaction and more directly so as to create a cloud of cells when
agitation of the sample container 43 occurs. A fluid having a
predetermined density and viscosity is not required, as in the
instance of centrifugation, in order to create mixing. That is to
say, mixing can be sufficiently accomplished, for example, through
vertical (e.g., up and down) movement of the sample container 43
causing the mixture (not shown) to move, arrows 61, 63 between the
adjacent chambers 51, 53, 55 and the transition zones 44, 48
therebetween of the sample container and producing the
agglutination reaction as well as the moving field of cells.
Depending on the volumes involved, however, the movement could be
made at an angle other than vertical. Alternatively, pumps or other
aggressive liquid moving means capable of moving a quantity of
liquid sample between the adjacent chambers for mixing can be used
to draw the fluid between the chambers in a similar manner. For
example, a single pump can be used to draw the liquid upwardly into
the chambers 51, 53 from chamber 55 wherein the force of gravity
will cause the moving flow field to move past the measurement
window. The sample container, as well as the various movement
mechanisms that can be used in conjunction therewith, are described
in greater detail in U.S. Publication Numbers US 2002/0076826 and
US 2002/0081747, each of which were previously incorporated above
by reference in their entirety.
[0054] Referring to FIG. 6, a second preferred form of test
chamber/testing system relies chiefly upon use of centrifugation as
the mechanism for transporting the reaction mixture to the reaction
and detection zone. According to one working example, a centrifuge,
shown only partially as 74, is used to support a plurality of
tubular sample containers 70 into which patient sample is metered.
The sample containers are generally as those described with regard
to FIG. 1, with some notable exceptions as described in greater
detail below. Preferably, each sample container 70 is a tube that
includes a planar measurement window such as shown in FIG. 5, to
permit detection. Reactant material (e.g., red blood cells) is
preferably contained in an upper portion of the container and a
receiving fluid having a specified density and viscosity is
provided in a lower portion (not shown) of the container. This
material can include at least one liquid having a higher specific
gravity than that of the blood cells. The other properties herein
contained of this at least one additional fluid are designed to
produce an environment that maintains the integrity of the red
blood cells.
[0055] As noted and though the present method specifically
describes agglutination reactions with regard to blood typing, it
should be apparent to one of sufficient skill and as described in
greater detail below that there is potential to use antigen
carriers other than red blood cells in that the herein described
particle detection system does not depend on the red color of the
cells.
[0056] First and with regard to centrifugation, reference is now
made to FIGS. 7-10, herein describing a proposed testing procedure
for blood antibody detection (FIGS. 7 and 8) and an proposed
testing procedure for A-B-O blood typing (FIGS. 9, 10), each using
the presently described method.
[0057] As noted, the sample containers 80 used in each of the above
tests are somewhat similar to those described with regard to FIG.
1. However, the restriction orifice 88 located between the upper
portion 84 of the container and the lower potion 86 thereof is
optional since spatial separation is not the preferred mode by
which the cells are classified following reaction. Preferably,
however, a restriction orifice is added that promotes a dispersion
of cells into a moving flow field. This restriction orifice,
however, is sized to be smaller than those of previously known
systems requiring both chemical separation and spatial separation.
The restriction orifice for this embodiment for example is
approximately 1 millimeter as opposed to a 4 millimeter diameter
required in known containers.
[0058] A receiving fluid containing an agglutination reagent is
added to the lower portion of the sample container, either as
filled within a clinical analyzer (not shown) or sold as a
prefilled quantity, the agglutination reagent being mixed with the
receiving fluid to form a homogenous mixture.
[0059] In the upper portion 84 of the sample container 80, a
patient serum is first added, followed by a RBC (red blood cell) or
other suitable reagent, the serum and reagent being added
preferably by means of a metering mechanism or pipette tip (not
shown). The edges of the restriction orifice 88 provide a latch
point for the metered serum material, thereby forming an air bubble
90 between the gel/reagent mixture and the serum/RBC reagent prior
to the application of the centrifugal force.
[0060] As to the constitution of the receiving fluid, a sugar-based
or other material having a density chosen to permit the red blood
cells to move through it at a specified rate under centrifugal
force is preferred.
[0061] According to an alternate embodiment, as shown in FIG. 7(a),
the sample container 80 can include two or more materials
designated 94 and 96, respectively having differing material
(density) properties. Preferably, a higher density material can be
provided in the lowest portion of the container while a lower
density material can be provided in an intermediate portion of the
container. According to this alternate embodiment, the red blood
cells would not settle at the bottom of the test container, but
rather would cease migrating when they reach a neutral buoyancy
rate. The latter is less sensitive to centrifugation, force and
time and therefore may enable more rapid centrifugation of the
sample container.
[0062] Still referring to FIGS. 7 and 8, the sample container 80 is
incubated wherein the bound antibody is bound to the red blood
cells. The sample container 80 is then centrifuged and test spun in
order to group the cells and force same through the restriction
orifice 88 where the cells are dispensed into a moving flow field
into the reagent matrix below. The centrifuge promotes the
agglutination reaction vis a vis the contained reagent. The moving
cell field is pushed as a "cloud" into the measurement windows 87,
89 of the container 80 wherein the particle detection system 32, 36
determines the size distribution of particles in order to determine
the strength of agglutination.
[0063] The above read process can occur in several ways: First, the
read can occur during centrifugation if synchronized with the
rotation of the centrifuge. This read technique provides real time
data relating to the state of the cells moving through the media,
thereby making the measurement insensitive to the final position
and minimizing the time to result, especially for strong
agglutination reactions.
[0064] Alternately, the read process can also occur after the
completion of centrifugation, but can proceed while the container
(e.g. tube) is still within the centrifuge. This enables
consolidation of hardware and uses the centrifuge as the location
device that holds the tube(s) in fixed position. In the embodiment
shown, the light emitter 32 and detector 36 of the light
scattering/particle detection system are arranged in a fixed
location relative to the centrifuge, as the centrifuge rotates the
tubes therebetween, per arrow 76. Each of the above read process
steps can therefore be accomplished using this type of system
station positioning.
[0065] Alternatively, the above read/detection process can also be
performed as a separate or off line procedure in which the tubes 70
can be placed in a separate device/apparatus (not shown) following
the centrifugation process. This latter process may be required if
there are space/size or other unique requirements that are not
available internally to the centrifuge.
[0066] Therefore, the sample container can be removed or remain in
the centrifuge for either an end point measurement or alternately
an "on the fly" measurement can be made in which at least one image
can be obtained while the centrifuge is slowing down or through use
of a strobe.
[0067] Alternative centrifugation techniques, for example,
involving a fixed angled centrifuge can be similarly utilized. This
technique has been shown to create a "smear" of cells along the
wall of the tubular sample container. A high-resolution vision
system or a low-angle particle analysis system such as that shown
in FIGS. 3 and 4 can then inspect these cells. As with a swinging
bucket centrifuge, it is possible to read the test tubes in the
centrifuge, during or after centrifugation, or after the tubes have
been removed from the centrifuge in a separate apparatus.
[0068] Referring to FIGS. 9 and 10, a similar sample container 80
is shown for A-B-O blood typing preferably having a smaller
restriction orifice, as compared to those of the prior art, for
example FIG. 1, in order to promote disposition of a moving flow
field. According to the technique, whole blood and a liquid serum
reagent are each added to the upper portion 84 of a sample
container 80. A gel is added to the bottom portion 86 of the sample
container 80, the gel having a density and viscosity that is lower
than that of the agglutinated red blood cells to promote movement
therethrough as a flow field.
[0069] The sample container 80 is incubated sufficiently and
centrifugation forces intimation of the cells, enhancing the
agglutination reaction as the cells are dispensed through the
restriction orifice 88 into the gel matrix and past the measurement
windows 87, 89 permitting scanning by the detection system 32,
36.
[0070] Using a test chamber set-up as shown in FIG. 5, an A-B-O
blood typing procedure/testing is performed by aspirating whole
blood and serum reagent into the confines of the container and then
promoting movement between the transition zones of the tip, thereby
creating the agglutination reaction. An air bubble interface
between the compartments of the container is optional for this type
of testing.
[0071] As previously noted, other forms of classification detection
can be performed using the above-described methodology.
[0072] Antibody detection can also be performed using a sample
container 100 similar in design to that depicted according to FIG.
5. According to this technique, serum cell material is initially
aspirated into the container 100. RBC reagent is then additionally
aspirated into the container 100. The serum and the RBC reagent are
then mixed through means of vertical agitation using a pump or
other means in order to create bound antibodies. The sample
container 100 is then axially spun by centrifuge or other means,
the container preferably having a compartment (not shown) within
same into which the cells are trapped as described in U.S. Pat. No.
4,933,291 to Daiss et al., the entire contents of which are
incorporated by reference.
[0073] Surplus material (reagent, serum, unbound antibodies) is
then discharged from the sample container 100 and a wash fluid is
aspirated into the container to wash the cells wherein the wash
fluid and cells are suspended and the sample container is axially
spun. The wash fluid is then discarded and agglutination reagent is
then aspirated into the sample container. The cell/reagent mixture
is resuspended and mixed by vertical agitation to create an
agglutination reaction by means of the transition zones after which
the passing mixture is read using the particle detection system and
determining a size distribution of the passing particles.
[0074] Referring to FIG. 11, and in lieu of using a separate
compartment, and axial spin (e.g., centrifugation) a magnetic bead
or particle can be alternately attached to each cell/bound
antibody. The sample container 43 can then be placed within a
toroidal magnet 100, suspending the cells, permitting discharge of
surplus material and use of a wash fluid to first wash unbound
antibody from the cells and then adding an agglutination reagent,
the magnet being used to retain the bound cells in the container 43
between washes and other processing steps.
PARTS LIST FOR FIGS. 1-11
[0075] 10 cartridge [0076] 12 sample container [0077] 14 sample
container [0078] 16 sample container [0079] 18 tubular sample
container [0080] 19 band, agglutination [0081] 20 sample container
[0082] 21 reaction zone, initial [0083] 22 sample container [0084]
23 upper portion [0085] 25 lower portion [0086] 26 button of cells
[0087] 30 matrix of inert particles [0088] 32 light emitter [0089]
33 light beam [0090] 35 lens [0091] 36 light detector [0092] 37
lens [0093] 38 detector rings [0094] 41 processor [0095] 43 sample
container [0096] 44 transition zone [0097] 48 transition zone
[0098] 51 chamber [0099] 53 chamber [0100] 55 chamber [0101] 61
arrow [0102] 63 arrow [0103] 70 sample container [0104] 74
centrifuge [0105] 76 arrow [0106] 80 sample container [0107] 84
upper portion [0108] 86 lower portion [0109] 87 measurement window
[0110] 88 restriction orifice [0111] 89 measurement window [0112]
90 air bubble [0113] 92 gel material [0114] 94 gel material [0115]
100 magnet [0116] .PHI..sub.i beam power, incident [0117] VSF
volume scattering function [0118] .DELTA.r thickness, scattering
volume [0119] .DELTA..OMEGA. solid scattering angle [0120] .psi.
scattering or pick-up angle [0121] .PHI..sub.s scattered power of
part of incident beam
[0122] Though the invention has been described in terms of certain
embodiments, it will be readily apparent that there variations and
modifications that can be performed that still embody the inventive
scope of the invention.
* * * * *