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Description  |
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BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to devices for detecting the presence of
analytes. In particular, the invention relates to such devices whereby
disposable assays may be quickly and efficiently conducted in the field.
More particularly, the invention relates to an assay device which may be
selectively controlled by an actuation device of predetermined speed
and/or pressure.
2. The Prior Art
There is a present and continuing need to detect a wide variety of analytes
with high specificity and high sensitivity in many applications. A
technique that is well known in the art uses antibody/antigen (antibody
generator) reactions to determine a target analyte. One common use of the
antibody/antigen pair is in the construction of a reaction environment in
which microscopic particles to which antibody or antigens have been
chemically attached are made to agglutinate or are inhibited from
agglutinating in the presence of the mating antibody/antigen and the
target analyte.
When an agglutination reaction occurs, the microscopic particles chemically
bind to each other with the antibody/antigen molecules serving as very
specific chemical binding agents, forming much larger aggregates of
particles which can grow in size to become visible to the naked eye. The
progress of the reaction may be monitored and resulting data analyzed to
provide quantitative and qualitative results on target analyte
concentration.
A specific agglutination reaction is latex agglutination. Latex
agglutination tests are available which detect small qualities of antigen
molecules. Agglutination reactions usually involve the aggregation of
latex particles which bear on the surface antigenic molecules. Aggregation
(agglutination) occurs when antibody molecules specifically corresponding
to the antigen (e.g. cocaine) are introduced into the solution of the
carrier particles. Antibodies can be visualized as having a "Y" shape
where both arms of the "Y" can attach antigen. Mixing antigen-coated latex
particles and antibody causes these components to interact and combine. As
more antibodies and particles become involved, many cross-linkages are
formed and the particles group together as visible clusters. However, when
free, unbounded antigen is introduced from an external sample, for
instance, agglutination does not occur. The free antigen caps the antibody
binding sites and inhibits the agglutination.
Devices are known in the art which can detect various analytes. However,
these devices require numerous steps that are not conducive to being used
in the field environment, with minimal training, in a simple sequence, to
provide consistent results. An assay device designed for ease of use by
the person performing the test would be desirable.
Devices are known which require a user to add the required reagents and
preform a crude stirring or mixing operation which is subject to not being
repeatable in the field. Carrying the required reagents, measuring the
exact proportions, and mixing the reagents in the assay device are steps
which are not conducive to ease of use with minimal training. It would be
desirable to have an assay device which contained the premeasured reagents
needed to perform a specific test to determine if a suspect substance
contained a target analyte. Additionally, it would be desirable to have an
assay device which was designed to adequately mix the reagents upon being
actuated before being introduced to the sample or suspect substance for
further mixing.
Known devices require the assay device to be held still or horizontal while
the reagent and sample mixture flow through the assay device to get an
agglutination result. Some devices require close tolerances in
manufacturing to obtain capillary flow. Other known devices require the
white room environment of a laboratory. It would be desirable for
operators in the field to have an assay device that could be used by
simply placing a sample to be tested into the device and triggering an
actuator which would quickly and repeatedly carry out the test without
room for operator error. It would also be desirable to have a device
resistant to an abusive environment of rough handling and jostling around
even while the test was being conducted.
This application is related to U.S. Pat. No. 5,290,517 entitled "Optical
Agglutination Assay Device", filed Apr. 4, 1990 and assigned to the same
assignee as the present application. and merely incorporated by reference.
While the applications disclose different design configurations for the
assay device, both may be utilized with an optical transmitting and
receiving unit for measuring the intensity of light reflected from or
transmitted through an optical viewing area in a track as a measure of the
occurrence of agglutination in the reaction system.
A simple, inexpensive, portable, disposable, user friendly device for the
collection and subsequent reaction of an unknown analyte with the
requisite reagents is disclosed in this application.
SUMMARY OF THE INVENTION
Accordingly, it is an object of the present invention to provide an
apparatus for the detection and measurement of an analyte in a sample or
on a surface.
It is another object of the present invention to provide an apparatus for
receiving a sample, all necessary reagents and the requisite tracks for
controlling the mixing of the reagents and further mixing of the reagents
with the sample in a low cost disposable unit that provides for a simple
sample collection and user initiated reaction.
A further object of the present invention is to provide a unique collection
apparatus which will mate with a reusable photometric reaction cell
reading device and provide a qualitative or quantitative determination of
the analyte concentration in the test sample.
It is an advantage of the present invention that the reagents required for
testing a specific analyte are contained in the device and ready for
mixing with a suspect sample. The user simply need introduce the suspect
sample to the assay device, then actuate the device to cause the reagents
and sample to mix, whereby results can then be analyzed.
Additional objects and advantages of the invention will be set forth in the
description which follows, and in part will be apparent from the
description, or may be learned by practice of the invention. The objects
and advantages of the invention may be realized and obtained by means of
the methods, instrumentalities and combinations particularly pointed out
in the appended claims.
To achieve the foregoing objects, and in accordance with the purposes of
the invention as embodied and broadly described herein, an appropriately
designed assay device is provided in which an antibody/antigen
agglutination reaction may be initiated and optically analyzed. The
optical analysis can be either by the naked eye or a signal generated by a
photodetector to determine either qualitative or quantitative information
about the concentration of specific analytes which are present in the
reaction mixture undergoing agglutination. The techniques used in the
antibody/antigen interaction are specific and sensitive. Therefore, this
approach becomes a generic one for the detection of analytes for which an
antibody/antigen pair can be made. Furthermore, because of the unique
design of the assay device the apparatus can be easily configured into a
low cost disposable reaction assay device collector assembly for use in a
hand-held portable detection unit.
In accordance with the present invention, the assay device for analyzing a
sample of unknown substance by mixing a reagent with the sample comprises
a housing having at least one storage reservoir containing reagent. The
housing has an entry port for receiving the sample suspected of containing
a target analyte into the housing. At least one track for controlling the
reagent flow rate and mixing characteristics fluidly connects the reagent
with the entry port for mixing the reagents with the sample. The assay
device may further contain a track fluidly connected to the entry port for
reacting the reagent and sample mixture. The reacting track may have a
viewing area for analyzing the results of the reagent and sample mixture.
One embodiment of the assay device in accordance with the present invention
includes a card having a first and opposing surface with an entry port
extending from the first surface to the opposing surface. A flexible
member contacts the opposing surface of the card to define at least one
storage reservoir. At least one track for controlling the reagent flow
rate and mixing characteristics is defined by the flexible member and the
card through which reagent can be transferred between the storage
reservoir and the entry port. The flexible member is deformable to force
reagent between the storage reservoir and the entry port. The device
further includes a track which is fluidly connected to the entry port for
reacting the reagent and sample mixture. The reacting track may have an
optical viewing area for viewing the results of mixing the reagent and the
sample for determining the sample substance identity. A delivery means for
conveying a sample suspected of containing a target analyte into the entry
port may be included. The device may additionally include mixing and
delivery means for transferring the reagent at a first flow rate to the
sample in the entry port for further mixing. The reagent and sample
mixture is then forced through the reacting track at a second flow rate
less than the first flow rate so that reaction can occur.
BRIEF DESCRIPTION OF THE DRAWINGS
The accompanying drawings, which are incorporated in and constitute a part
of the specification, illustrate a presently preferred embodiment of the
invention which, taken with the general description given above and the
detailed description of the preferred embodiment given below, serve to
explain the principles of the invention. In the drawings:
FIG. 1 is an exploded view of a preferred assay device which embodies the
concepts and principles of the present invention;
FIG. 2 is a top view of the preferred assay device shown in FIG. 1 without
a cover member;
FIG. 3 is a cross-sectional view taken essentially along the line 3--3 of
FIG. 2 showing a storage reservoir, controlling track, barrier and
yielding member prior to actuation;
FIG. 4 is a view as in FIG. 3 illustrating a storage reservoir, controlling
track, barrier and yielding member during actuation;
FIG. 5 is a top view of the preferred assay device with an engaged swab;
FIG. 6 is a side view of a preferred swab with protective closure in an
opened position;
FIG. 7 is a cross-sectional side view of the preferred swab shown in FIG. 6
in a closed position;
FIG. 8 is a cross-sectional side view of a preferred hinged swab in the
opened position;
FIG. 9 is a cross-sectional side view of the preferred hinged swab shown in
FIG. 8 engaged with a swab retainer strip;
FIG. 10 is a side view of the preferred actuation mechanism.
FIG. 11 is an exploded view of the assay device and swab in use with the
actuator and electronics which evaluate the test results; and
FIG. 12 is a schematic of the assay device in use with the electronics
which evaluate the test results.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
A preferred embodiment of the assay device 10 in accordance with the
present invention is illustrated in FIGS. 1 and 2. FIG. 1 shows an
exploded view of the device 10, while a top view of the device 10 without
a cover member 12 is illustrated in FIG. 2. Device 10 includes a housing
14, storage reservoirs 16 containing reagent, and track 18. The track 18
which is fluidly connected to the storage reservoirs 16 serves different
purposes at various stages of the track 18. The track 18 can be viewed as
having the following portions: controlling track 20 fluidly connected to
the storage reservoirs 16 for controlling the reagent flow rate and mixing
characteristics; receiving track or an entry port 22 fluidly connected to
the controlling track 20 for receiving a sample suspected of containing a
target analyte into the housing 14; reacting track 24 fluidly connected to
the entry port 22 for reacting the reagent and sample mixture; and an
accumulator track or accumulator reservoir 26 in the reacting track 24 for
retaining excess reagent and sample mixture.
The number of storage reservoirs 16 is dependent upon the reagents required
for detecting a particular target analyte. At least one storage reservoir
16 is needed to contain a reagent or mixture of reagents. If a particular
set of reagents can be premixed instead of being mixed just prior to
actuation with the sample then one storage reservoir may be all that is
required.
The controlling track 20 may have any number of fluidly connected paths to
the reservoir 16 for dividing, agitating and mixing the reagents. By way
of example and as best illustrated in FIG. 2, the controlling track 20
includes a first portion 20a, 20a' and 20a", each of a predetermined
cross-sectional area, in fluid communication with a respective one of the
reservoirs 16. Each portion 20a, 20a' and 20a" discharges into a
respective second portion 20b, 20b' and 20b", each of a cross-sectional
area greater than that of the first portion to which it is connected.
A third portion of controlling track 20 is formed by branches 20c and 20c'
which then joins with respective fourth portions 20d and 20d'. These
fourth portions 20d and 20d' both discharge into a fifth portion 20e of
controlling track 20. Fluid in fifth portion 20e is divided in sixth
portions 20f and 20f' which bend around to form final and seventh portions
20g and 20g' positioned to discharge fluid tangentially into the entry
port 22. Under circumstances where only one storage reservoir 16 is
required, the controlling track 20 may still provide a means for mixing
the reagent prior to introducing the reagent to the sample, thereby
eliminating any need for shaking the reagent prior to actuation. A nozzle
or set of nozzles 28 for assisting in mixing the reagents through changing
flow rates and turbulently mixing the reagents together are be included in
portions 20b, 20b' and 20b" of controlling track 20. Additionally, the
nozzles 28 can be used to control the flow rate of the reagent to insure
that the reagent flow rate is sufficient to mix the reagent and sample
together.
The nozzles 28 serve as a means for mixing the reagents by having the
reagents collide at an accelerated speed. In a preferred embodiment the
nozzles 28 are molded into the portions 20b, 20b' and 20b" of controlling
track 20 to form mixing points. Between nozzles 28, the cross-section of
the controlling track 20 may be increased in order to increase the
hydraulic diameter of the controlling track 20, which in turn lowers the
frictional head loss in the controlling track 20. Lower head loss will
allow more actuation energy used to force reagent from the reservoirs 16
to be used to increase fluid velocity instead of frictional losses. The
nozzles 28 may be created by stepping the controlling track 20 up or down
in cross-section. However, the preferred method is ramping the controlling
track 20 with smooth, straight or curved tapers. Not only does this method
lower the frictional head losses, but results in fewer air bubbles
entering the system. A preferred embodiment of controlling track 20 has
the reagent flow turn ninety degrees into portions 20c and 20c' after a
head-on collision of the reagents. After the reagents collide and are
turned to continue further down the controlling track 20, the controlling
track 20 is slightly larger in cross-sectional area so that no unnecessary
flow resistance is created. The number of nozzles 28 used in a particular
assay device 10 may vary with the specific reagent flow rate and mixing
characteristic requirements.
A preferred embodiment of the assay device 10 for detecting a target
analyte can be best understood by the exploded view of the device 10
illustrated in FIG. 1. The optical agglutination assay device 10 includes
a card 30 having a first surface 32 and an opposing surface 34 with an
entry port 22 extending from the first surface 32 to the opposing surface
34. A flexible member 36 contacts the opposing surface 34 of the card 30
forming at least one reservoir 16 containing reagent and covering the
track 18, which is molded into the opposing side 34 of the card 30. At
least one controlling track 20 for controlling the reagent flow rate and
mixing characteristics is defined by the flexible member 36 and the card
30 through which the reagent can be transferred between the reservoir 16
and the entry port 22. The flexible member 36 is deformable to allow the
reagent to be forced between the reservoir 16 and the entry port 22. The
reacting track 24 is fluidly connected to the entry port 22 for reacting
the reagent and sample mixture. The reacting track 24 has an optical
viewing area 38 for viewing the results of mixing the reagents and the
sample to determine the sample substance identity.
In a preferred embodiment the flexible member 36 covers the entry port 22
on the opposing surface 34 of the card 30 as well as the track 18 which is
preferably recessed into the opposing surface 34 of the card 30.
Additionally, in an alternative embodiment the card 30 and flexible member
36 may be composed of a single piece housing 14 having a flexible top
surface.
Preferably the card 30 is made of molded plastic, such as polystyrene. The
track 18 through which the reagent flows is molded directly into the card
30. The reservoirs 16 are vacuum-formed into a sheet of polyester film,
while the film is being heat-sealed over the side of the card 30 in which
the track 18 is molded. This straight-forward process takes less than a
minute to complete.
Fill holes 40 are used for filling the reagent storage reservoirs 16 with
reagent. Vent holes 42 allow excess air to be displaced from the system
during the filling process. Another ventilation hole 44 is shown at the
opposite end of track 18 from where the track 18 connects to the
reservoirs 16. The purpose of this ventilation hole 44 is to permit any
excess air to escape from the track 18 when the reagents are forced from
the reservoirs 16. Tape 46 covers the holes 40, 42, and 44 respectively.
The tape 46 covering ventilation hole 44 is preferably removed prior to
conducting a test with the device 10.
To prevent reaction product from leaking out of the ventilation hole 44 the
accumulator reservoir 26 is molded into the reacting track 24. This cavity
has the capacity to hold all of the reagent volume in the reservoirs 16.
In a preferred embodiment a cover member 12 made of molded plastic is used
to sandwich the flexible member 36 between the card 30 and the cover
member 12 to protect the system from being ruptured. Additionally, the
cover member 12 has an opening 48 to allow access to the reservoirs 16 for
actuating the device 10 by applying pressure to the reservoirs 16 to force
the reagents through the track 18. The reservoirs 16, which are not as
high as the cover member 12 is thick, protrudes into the opening 48. The
optical viewing area 38 is in the cover member 12 to enable the reaction
of the reagents and sample or reaction product as it passes through the
reacting track 24 to be observed to determine the results of the test. The
cover member 12 is ultrasonically welded to the card 30; although the
cover member 12 could also be attached with adhesive or doubled-sided tape
by way of example.
A removable label 50 covers opening 48 in the cover member 12 to further
protect the reservoirs 16 from accidental actuation. Additionally, the
label 50 provides information identifying the target analyte the device 10
is designed for testing. A portion 51 the label 50 covering the reservoirs
16 and viewing area 38 is simply removed after the sample has been
introduced to the device 10 and the user desires to carry out the test.
The label 50 is made of stiff paper and coated with a high tack/low tack
adhesive so that it can be easily and cleanly removed by the user at the
appropriate time.
A means 52 for preventing the reagents from leaking from the reservoirs 16
due to jarring or dropping the device 10 is shown in FIGS. 1 and 2 and
illustrated in cross-sectional view in FIGS. 3 and 4. The preferred means
52 for preventing reagent leaking until actuated by an operator utilizes a
controlling track barrier 54 and a yielding member 56.
The barrier 54 is molded directly into the card 30 as shown in FIGS. 1-4.
The barrier 54 is essentially a break in the controlling track 20 leading
out of each reservoir 16. The top 58 of the barrier 54 is coplanar with
the opposing surface 34 of the card 30. The wall of the barrier 54 closest
to the reagents is preferably perpendicular to the bottom of the track 18
or has a reverse slope so that the fluid is not directed up over the
barrier 54. A platen, which is used to seal the polyester film to the card
30, contains a cavity directly above the barrier 54. The cavity covers a
larger area than the top 58 of the barrier 54. This cavity is vented to
the ambient atmosphere, so that the heated air in the cavity can escape
during sealing. If the cavity were not vented, the air within the cavity
would increase in pressure, thereby causing the film to seal to the top 58
of the barrier 54. When the heat sealing operation is completed, the heat
seal film ends up lying against the top 58 of the barrier 54, thus closing
off the passage to the controlling track 20. This prevents the reagents
from moving down the track 18 if the card 30 is jarred, shaken or dropped.
However, when pressure is applied to the reagents contained in the
reservoirs 16, such as occurs during actuation, the reagents will flow
through the track 18 and over the top 58 of the barrier 54 by deflecting
the yielding member 56, as illustrated in FIGS. 3 and 4.
The introduction of the barrier 54 into the track 18 also creates some
resistance to the reagent flow when the reagents are forced out of the
reservoirs 16. Some of the energy applied to the reservoirs 16 during
actuation is used to push the fluid up and over the barrier 54. A portion
of the energy is used to deform the yielding member 56.
The device 10 is intended for use with any reagents capable of utilization
in determining the identity of a target analyte where the ease,
convenience and reliability of having a self contained system are desired.
Specifically, in an agglutination reaction, it is known in the art that a
reaction between reagents and a target analyte may be designed to produce
agglutination or the inhibition of agglutination.
However, a preferred embodiment of the present invention would utilize a
latex agglutination reaction method for detection where antibodies are
able to recognize minute quantities of the substance of interest, for
example, cocaine. Latex agglutination reagents which detect various drugs
of abuse are commercially available from Roche Diagnostics, Nutley, N.J.
Each of the known test kits is packaged as a set of three reagents for
testing for cocaine, one of which is a solution of latex particles. For
optimal effectiveness, the latex particle solution must be shaken
immediately prior to use. This is recommended because the latex particles
have a density greater than the surrounding solution causing the latex to
settle over short periods of time.
It is preferred that the reagents used in the assay device 10 of the
present invention eliminate the need for shaking the latex particle
solution. The density of the latex particle solution is increased to equal
the density of the latex particles thereby preventing particle settling.
Roche Diagnostics provides a latex agglutination test kit for cocaine
detection containing: an antibody reagent A--one vial of mouse monoclonal
anti-cocaine antibody in a buffered solution; reaction buffer B--one vial
of buffer reagent; and latex reagent C--one vial of latex-cocaine
particles in a buffered solution.
The Roche Diagnostics test requires the sequential addition of one drop of
each reagent with an external or suspect sample. However, the procedure
recommends "invert(ing) reagent C approximately 8 to 10 times before use.
If excessive foaming is observed allow it to settle before using."
A preferred embodiment of the latex reagent would include the addition of
133 milligrams of sucrose in a final volume of 1 milliliter of latex
reagent C creating a latex particle solution having a solution density of
1.05 gram/milliliter, equaling the density of latex particles. As a
result, the latex particles remain suspended in the buffered solution and
the requirement for inversion of reagent C prior to use is eliminated.
To assess the effectiveness, samples were tested utilizing the reformulated
C reagent in the latex agglutination test for cocaine. Ten positive
samples (buffer containing 100 parts per million cocaine) and ten negative
samples (buffer containing 10 parts per billion cocaine (less than the
cut-off value) were tested. Each test produced the appropriate positive or
negative result.
This process can be used to enhance the effectiveness of other latex
agglutination tests for drug detection. For instance, latex agglutination
tests for morphine (heroin), phencyclidine (PCP), marijuana and
methamphetamine are commercially available in a similar design as the
cocaine test kit discussed above. Respective C reagents from each of these
latex agglutination tests can be modified as described above resulting in
an easier to use product.
In a preferred embodiment of the assay device 10 having three storage
reservoirs 16 for containing the cocaine detecting reagents discussed
above, the nozzles 28 at the head-on collision are not symmetrical. The
nozzle 28 for the center reservoir 16 is only tapered in one dimension,
instead of two. The point at which the collision occurs is not on the
centerline of the downstream portion (20c or 20c') of the controlling
track 20. The intersection is biased towards the center reservoir 16 so
that more of the outer reservoir 16 reagent flows through the
intersection. This is done because each of the three reservoirs 16 contain
the same amount of reagent, but the contents of the center reservoir 16 is
divided equally between the two initial collisions, i.e. one collision
with the reagents of the other two reservoirs 16.
The shape of the nozzles 28, as well as the location of the collision
point, work together to mix equal volumes of the reagent from the center
reservoir 16 with the reagents from the remaining two reservoirs 16. The
shape and size of the nozzle 28, along with the dimensions of the
controlling track 20 preceding the nozzle 28, i.e., the cross-sectional
areas of respective portion 20b, 20b' and 20b", can be adjusted based on
the properties of the fluids involved and the mixing ratios desired
A delivery means 60 for delivering the sample suspected of containing a
target analyte into the device 10 is illustrated in FIG. 5. A preferred
embodiment of the delivery means 60 for conveying the sample to the assay
device 10 is a swab 60. FIG. 5 shows a top view of a preferred assay
device 10 with the swab 60 engaged into the entry port 22. Swab 60 is used
to collect an unknown sample by rubbing it in the sample or by simply
moving it along a surface which is suspected of having been contaminated.
The device 10 works by mixing the reagents contained in reservoirs 16 with
the sample introduced to the system by the swab 60. After mixing, the
results may be determined by analyzing the solution in the reacting track
24.
The cylindrical interface between the preferred swab 60 and the entry port
22 is both easy to manufacture(i.e. mold) and easy to use. A sharp,
tapered ridge around the inside of the entry port 22 acts like the barb on
a fishhook, allowing the swab 60 to be easily pushed past the ridge in one
direction, while preventing it from easily being removed in the opposite
direction. The ridge digs into the surface of the swab 60, which is
preferably made of a soft plastic, to create a fluid seal.
The swab 60 illustrated in FIG. 6 is in the fully opened position, while
FIG. 7 illustrates the closed or protected position. To improve the
reliability of mixing the reagents with the sample, the gathering surface
62 of the swab 60 is covered with many closely spaced, small cones 64
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