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Description  |
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FIELD OF THE INVENTION
The present invention relates to methods and apparatus for controlling the
flow of fluids through a substrate such as a pad or filter, particularly
for use in clinical assay devices.
BACKGROUND OF THE INVENTION
The ability to measure a wide variety of physiologically active compounds,
both naturally occurring and synthetic, has become of increasing
importance, both as an adjunct to diagnosis and to therapy. While for the
most part, assays of physiological fluids and drugs have required clinical
laboratory determinations, there is an increasing awareness of the
importance of being able to carry out assay determinations in the
physician's office or in the home. To be able to perform an assay in a
physician's office or home requires that the assay have a simple protocol
and be relatively free of sensitivity to small changes in the conditions
under which the assay is carried out.
A wide range of disposable assay devices has been developed for use either
in analytical laboratories or in physicians' offices or homes. These
devices, because they are used by inexperienced operators, should be
simple to operate and should incorporate all the reagents necessary for
the test to be conducted.
One analyte of importance is cholesterol. There is a clearly established
relationship between total blood cholesterol (mainly the LDL fraction) and
coronary artery disease (J.A.M.A. 253: 2080-2086, 1985). New guidelines
have been established for adults to identify risk groups associated with
blood cholesterol levels. Since cholesterol levels can be controlled by
both diet and cholesterol-lowering medications, it is useful for those
individuals at risk to be able to monitor their own cholesterol at home in
order to reduce the potential for heart disease. The measurement of other
naturally occurring compounds of physiological importance, such as
glucose, lipoproteins, etc., as well as synthetic drugs, is also of great
interest. For example, therapeutics, drugs of abuse, iodothyronines,
alcohol, cytokines, as well as numerous other chemical analytes could be
monitored. Also of interest are microorganisms, .beta.-HCG for ectopic
births, antibodies associated with disease, and the like.
Many of the most commonly used assays in disposable assay devices require
an incubation step, such as requiring enzymes to act on the sample, such
as for determinations of cholesterol, glucose, uric acid, and the like.
Additionally, enzymes are often used as labels in immunoassays. In a
conventional enzyme immunoassay, an enzyme is covalently conjugated with
one component of a specifically binding antigen-antibody pair, and the
resulting enzyme conjugate is reacted with a substrate to produce a signal
which is detected and measured. The signal generated by the enzyme, in
either the conventional chemical assay or the immunoassay, may be a color
change, and the color change may be detected with the naked eye or by a
spectrophotometric technique.
Many of the disposable assay devices currently in use include one or more
reagent zones comprising layers incorporated with assay reagents. Among
the problems encountered in use of these devices is the premature
interaction or migration of these reagents, either during the
manufacturing process or upon introduction of the sample to the device.
Both enzymatic and chemical reactions often require incubation steps. One
of the challenges in designing a truly "one-step" disposable device is to
provide a means to delay the fluid flow in order to allow for proper
incubation periods. This is particularly challenging for non-instrumented
disposable analytical devices.
Ideally, a disposable assay device should include a means to delay the flow
of the sample through the device for a predetermined time to permit
incubation of the sample with the reagents or indicators present in a
particular region of the device. After the incubation period, which is
generally on the order of a few minutes or less, the sample then flows to
the next region of the device for further processing.
An ideal flow delay means should work like a valve, with a "closed" and an
"open" state. When the state is "closed", the fluid flow should stop, and
when the state switches to "open", the fluid should flow through the
flow-delay valve with little or no restriction, and the flow rate of the
fluid through the device should be unchanged.
A wide range of disposable analytical devices has been developed which
include means to control flow of fluids therethrough. However, none of
these previously developed devices has a flow-delay means with a
valve-like effect on the flow of fluids.
Deutsch et al., in U.S. Pat. No. 4,522,923, disclose a test device
comprising a container having at least two water-soluble barriers between
at least three superposed chambers. Upon introduction of an aqueous
biological sample to be tested into the topmost chamber, the sample will
successively mix with the contents of the chambers. The contact time in
each chamber is a function of the water solubility of the barriers.
Ebersole, in U.S. Pat. No. 4,522,786, discloses a multilayer test device
comprising at least two liquid permeable functional layers superposed upon
one another, the layers in liquid communication, with a barrier layer
separating the layers. The barrier layer is a chemically inert, liquid
insoluble, foraminous septum, the foramina of which are filled with a
thermally sensitive material which is liquid impermeable at assay
temperatures but capable of melting when heated to provide rapid liquid
communication between the function layers. This melted material then
travels through to the next layer with the test liquid.
Jones, in U.S. Pat. No. 5,213,965, discloses an assay device for measuring
high density lipoprotein or cholesterol in a fluid sample which contains
other lipoproteins. Sieving materials chromatographically separate
aggregated from non-aggregated materials in the sample as the sample flows
through the matrix. A reagent reservoir slowly releases a precipitating
agent into the matrix by formulating the precipitating agent with a binder
for slow dissolution on contact with a sample. There is no provision for
timing the delay of the sample within a region.
Vonk, in U.S. Pat. No. 5,185,127, discloses a device for assaying an
analyte comprising an enclosure and a filter stack. Flow control is
provided by a hydrophilic membrane which contains a binder for an analyte.
This membrane is impervious to the passage of an aqueous liquid until
activated by a wetting agent.
Johnston et al., in U.S. Pat. No. 4,038,485, disclose a test composition
for detecting a component in a sample which comprises a reactant system
which, upon contact with the sample, interacts with the component to
produce a detectable response, as well as an inhibitor system which, upon
contact with the sample, prevents the reactant system from interacting
with the component after lapse of a predetermined time.
Amano et al., in U.S. Pat. No. 4,889,797, disclose a dry analytical element
for assaying enzyme activity in a liquid comprising a support having
provided thereon at least a porous liquid-spreading layer composed of
fibers which do not absorb water. Here, flow control is directed to
decreasing spreading of the liquid within a layer rather than through a
layer.
Deneke et al., in U.S. Pat. No. 4,876,076, disclose an assay device
comprising a first carrier layer having applied thereto a liquid absorbing
layer and a separate second movable carrier having applied thereto a
dissolvable reagent-containing layer which is not in initial contact with
the first carrier layer. The reagent-containing layer is dissolved by
contact with a liquid contained in the liquid absorbing layer, and the
first carrier layer is positioned in the device to permit contact between
the liquid absorbing layer and the reagent-containing layer by applying
pressure to one of the carrier layers. Flow of sample from one layer to
another is controlled by bringing the two carrier layers into contact with
each other so that the sample is transferred. This type of flow control is
also shown in Ramel et al., U.S. Pat. No. 4,959,324, in which a flow path
is completed by moving a sample receiving pad into a gap between two assay
strips.
Woodrum, in U.S. Pat. No. 4,959,305, discloses a multizone test device for
immunoassays in which assay reagents are reversibly immobilized within the
various layers of the device. The binding interactions within the device
depend upon the interactive properties of and between the assay reagents
and the matrix comprising the incorporating layer of the reversibly
immobilized assay reagents, and the disruptive properties of the liquid
test sample necessary to disrupt the particular reversible binding
interaction between the assay reagents and the matrix to release and
render useable the reagents in an analytically effective amount within the
device.
Columbus, in U.S. Pat. No. 4,549,952, discloses a liquid transport device
having means for increasing the viscosity of the liquid when the liquid
flows past at least one surface of the device. Control of flow is achieved
strictly through viscosity increase.
Hydration and expansion of a compressed foam switching element permit
automatic timed sequential delivery of multiple reagents in a device
disclosed by Bruce et al. in Analytical Chemical Acta 249 (1991), 263-269.
This technique is not well-suited for devices that require no diluent and
which are sensitive to the total volume of sample fluid required.
Hillmann et al., in U.S. Pat. No. 4,963,498, disclose methods for using
specific binding pair members which result in agglutination formation. The
resulting agglutinated particles may provide for changes in flow rate.
Interrupting capillary flow of liquid between two pieces of bibulous
material using a liquid expandable material is shown in Kurn et al., U.S.
Pat. No. 5,104,812.
Physical barriers to reduce the flow rate in a zone of a liquid transport
zone are shown in Columbus, U.S. Pat. No. 4,310,399; Columbus, U.S. Pat.
No. 4,618,476; and Grenner et al., in U.S. Pat. No. 5,051,237.
Liquid flow through a filter can also be controlled by reactions between
the sample and a component of the filter, cf. U.S. Pat. No. 5,217,905, to
Marchand et al. Similarly, Tanaka et al., in U.S. Pat. No. 4,966,784,
disclose inhibiting migration of a water-soluble indicator in a reagent
layer in an assay device by using a particular organic solvent.
Liotta, in U.S. Pat. No. 3,723,064, discloses a layered testing device
including a first porous layer impregnated with a reagent system which
reacts with the analyte to produce an end product. A membrane having
plural regions with differing permeabilities is adjacent to the first
porous layer. The permeability differences are obtained either by
impregnating the regions with different concentrations of a chemical
reactive with the end product, or by varying the pore size in the regions.
Engelmann, in U.S. Pat. No. 4,738,823, discloses a test strip which has a
preselected capacity for absorbing sample. In this case, however, the
amount of sample applied to the test strip is metered, rather than the
amount of sample fluid that is retained for a predetermined time within a
selected area of a test device.
Other workers have coated filters with a variety of coatings to alter
conditions within the filter, none of which provides a valve-like action
to control the flow of fluid: Nagatomo et al., U.S. Pat. No. 4,587,102.
A layer for controlling diffusion rate of sample through the device can be
provided by varying the formulation ratio of a hydrophobic polymer and a
hydrophilic polymer which constitute the layer, as shown by Koyama et al.,
U.S. Pat. No. 4,615,983. In a similar fashion, Rothe et al., in U.S. Pat.
No. 4,587,099, disclose a test strip which includes a slowly absorbent
layer for sucking up a fluid sample. This layer is slowly absorbent rather
than a layer which retains a liquid for a predetermined period of time.
A flow-delaying polymer on an immunoassay filter is shown in Nelson, U.S.
Pat. No. 4,923,680.
Blood clotting has been used to inhibit sample flow through a track in
Lucas, U.S. Pat. No. 5,207,988.
Leeder et al., in U.S. Pat. Nos. 4,837,395 and 5,089,383, disclose a
heterogeneous immunoassay in which the production of the signal is
temporarily delayed by using an inhibitor which can be an alternate
substrate for the enzyme signal or a compound which reacts with the
product of the enzyme and its substrate.
Terminiello et al., in U.S. Pat. No. 4,774,192, disclose a dry chemistry
reagent system comprising a porous membrane of essentially uniform
composition which has a porosity gradient from one planar surface thereof
to the other.
Kuroda et al., in U.S. Pat. No. 4,416,777, disclose a material for
separating leukocytes from a leukocyte-containing suspension which
comprises a fibrous material having a surface layer coated on the fibrous
material which can be dissolved by degrees in water.
None of the above-noted patents provides a reliable means for metering the
rate of flow delay through a layer in order to retain a sample in contact
with a reagent for a predetermined length of time, nor where the length of
time can be varied depending on the incubation time required by the assay.
Alkyl ketene dimers have been used for many years as reactive alkaline
sizing agents in the paper industry. Industrially, alkyl ketene dimers are
added to the "wet end" of the papermaking process, that is, the slurry of
bleached pulp in water that is at the very start of the papermaking
process. Aqueous alkyl ketene dimer emulsions, which contain alkyl ketene
dimer and additives such as defoamers and biocides, have been developed
and marketed for this application. Wet-end addition allows for th | | |
