Flow through assay apparatus

Abstract: Assay apparatus (10) is provided which includes a filter (24) such as a porous membrane, a volume (18) that can contain an absorbent and flexible means (20) for reducing the volume (18) in order to effect passage of liquid sample through the membrane (24) and into the volume (18). The assay apparatus can be produced by a continuous manufacturing process.

Description: BACKGROUND OF THE INVENTION This invention relates to an apparatus for assaying a fluid sample to detect the presence or absence of a chemical or biochemical constitutent in the sample or in the surrounding environment. At the present time fluid samples are tested in a variety of apparatus for various constituents including specifically reactive pairs such as antibody-antigen pairs, enzyme-substrate pairs, D.N.A. fragment pairs, R.N.A. fragment pairs and the like. It is common practice to immobilize a reactant for the constitutent being tested on a support surface and then contacting the fluid sample with the coreactant. The constitutent is allowed to react with the coreactant to determine the presence, and, in some cases, the concentration of the constituent in the sample. Examples of assays which permit determination of the constituent concentration include the enzyme-linked immunosorbent assay (E.L.I.S.A.) and the radioimmunoassay (R.I.A.) In these procedures, filter materials such as microporous or ultrafiltration membrane, nonwovens, papers, glass fibers, or other filter matertials membranes provide a desirable immobilizing support for biologically active constituents since separation of the constituent for which the test is conducted can be easily effected. Examples of suitable membranes are disclosed in U.S.-A-4,066,512. Suitable apparatus utilizing these and other membranes to effect fluid sample assays are descrived in US-A-4,246,339; US-A-4,526,690; US-A-4,632,901; US-A-4,727,019 and US-A-4,737,192. While each of these apparatus provides the capability of conducting accurate fluid assays, each of these apparatus suffer from the same deficiencies. The primary deficiency is that they comprise multipiece constructions which require that the apparatus pieces be formed in a batch mode of construction such as by injection molding. Thereafter the pieces are assembled to form the final apparatus. Due to their design and method of assembly, these apparatus are expensive to produce. Accordingly, it would be desirable to provide an apparatus for effecting assays which can be mass produced without the need for manual labor. Furthermore, it would be desirable to provide such an apparatus which can effect either immediate reaction or permit an incubation period for the reactants. In addition, it would be desirable to provide such an apparatus which renders it possible to effect a multistep reaction process with variable incubation times being permitted for each step. Furthermore, it would be desirable to provide such an apparatus which can include one or a plurality of discrete reaction zones. Such apparatus would provide a desired economy in production and a desired flexibility in use of assay apparatus. From EP-A-0 272 044 there is known a diagnostic apparatus comprising an absorbent hydrophilic pad in a chamber the upper end of which is closed by a porous flexible member disposed such that a substantially constant distance is maintained between the absorbent pad and the flexible member. The purpose of the porous flexible member is to equilibrate the pressure in a portion of the apparatus which is subjected to a vacuum force and in a portion between a membrane filter and the absorbent pad so that liquid on the filter can be forced through the membrane under the vacuum force generated by reduced internal pressure. Thus, said reference does not contemplate the use of any driving force other than vacuum to effect filtration. From EP-A-0 353 025 there is known an apparatus for conducting an assay for a constituent in a fluid sample which comprises a membrane filter which during no-flow conditions is always in intimate contact with a non-absorbent material. When it is desired to activate the assay by having the sample flow through the membrane filter, the latter is slid from the non-absorbent material to contact the surface of an absorbent hydrophylic material so that passage of the fluid sample through said membrane filter is promoted. The present invention provides an apparatus as defined in the claim. In an apparatus according to the invention there is always a space between the membrane filter and the absorbent hydrophilic material during no-flow conditions. When flow or passage of the fluid sample through the membrane filter is desired, a compressive force is used to contact the absorbent material with the membrane filter by which the passage of the fluid sample through the membrane filter is created. BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 is a side view of one assay apparatus of this invention. Fig. 2 illustrates the use of the apparatus of Fig. 1. Fig. 3 illustrates a process for making the apparatus of Fig. 1. The present invention provides an assay apparatus and process capable of conducting chemical or biochemical assays such as ELISA or RIA wherein reaction is effected rapidly or an incubation reaction period is utilized in one or multiple steps. The apparatus of this invention is moreover useful to effect an assay of discrete fluid samples or to assay the environment surrounding the apparatus. In one form of this invention, the membrane has bound to it a specific receptor such as an antibody, antigen, DNA strand, RNA strand, enzyme, enzyme substrate or the like which is specific for a complementary ligand in a manner well known in the art. Referring to Figs. 1 and 2, the assay apparatus 10 includes a flexible wall 12 which functions as a diaphragm formed integrally with side walls 14 which, in turn, are formed integrally with flanges 16. The flexible wall 12, side walls 14 and flanges 16 together form a well 18 to house an absorbent material 20. The flanges 16 are sealed about the entire periphery of the well 18 to a liquid impermeable member 22 such as a plastic film or an adhesive coated film so that the well 18 can be hermatically sealed. The liquid impermeable member 22 includes one or more hole so that a filter 24 can be sealed to the member 22. The flexible wall 12 can be made of any thin flexible material that can be flexed manually as described below such as polystyrene, aluminum foil or laminate. The flexible wall can be made from any thermoplastic material, or a component of multiple films as long as they are formable and can be hermatically sealed to member 22. As shown in Fig. 2, liquid sample 26 is added dropwise to contact the membrane 24. The membrane can contain, entrapped therein particles such as latex particles having their surface made first with a conjugate adapted to bind a desired reactant in the sample. As shown in Fig. 2, the membrane 24 is separated from absorbent 20 by space 28. When the absorbent 20 is so-positioned, there is no driving force for the liquid sample 26 to be passed through the filter 24. If desired, the wall 12 can be flexed toward the membrane 24 to effect contact with the undersurface of the membrane prior to applying the liquid sample 26 to the membrane 24. In this case, the liquid sample will pass through the membrane immediately under capillary action. After incubation, if desired, is complete, the wall 12 is flexed to the position 12a so as to effect passage of the sample 26 through the membrane 24 leaving a filtrate, a bound reactant, exposed on the surface of on within the membrane 24 which can be detected by any conventional means such as ELISA or RIA. When it is desired to utilize the apparatus 10 of Fig. 1 by the method illustrated in Fig. 2 to detect a constituent in the atmosphere surrounding the apparatus 10, an ampoule containing a chemical composite is included within the well 18. When the wall 12 is flexed toward membrane 24, the ampoule is broken and the chemical composition migrates through absorbent 20 through filter 24 and contacts the atmosphere 28. The chemical composition then changes characteristics such as color when contacted with a composition in the atmosphere to be detected such as oganics such as ethizlene oxide or formaldehyde metal vapors such as mercury or gases such as carbon monoxide or nitrogen oxides. Reaction product in the membrane, if any, can be shown by any suitable technique. For use with E.L.I.S.A. procedures when the reagent and co-reactant are a specific antibody-antigen pair, a conjugate of an active enzyme with a biological material specific to the harvested reagent can be passed through the membrane, followed by washing as above described. A substrate-chromogen mixture specific to the enzyme is then passed through the membrane to develop visible color in the presence of harvested reagent. Preferably the test is performed with positive and negative controls in which control samples, (a) known to contain the reagent (positive) and (b) known to be free of reagent (negative), are passed through adjacent test wells and the control membranes compared with the test membrane(s). Referring to Fig. 3, the apparatus 10 of Fig. 1 is formed continuously by passing a flexible thermoplastic film 30 from roller 32 through a conventional vacuum or pressure forming step 34 to produce a continuous sheet 36 having a plurality of wells 18. Each of the wells 18 is filled with an absorbent 20 and the wells 18 containing the absorbent 20 is contacted with a porous filter 24 from roller 38 which is sealed in the flanges 16 of wells 18. A liquid impermeable cover 40 from roll 42 such as an aluminum foil laminated to a thermoplastic polymer is then heat sealed to the membrane 24 over the flanges 16. The cover 40 has holes spaced apart therein so that each assay apparatus 10 has a membrane 24 exposed on the top surface of the apparatus 10. Individual apparatus 10 then are formed by cutting the continuous sheet with conventional cutting apparatus such as knives 44.

Claims: Apparatus for conducting an assay for a constituent in a fluid sample which comprises: a membrane filter (24); a support (22) sealed to a first surface of said membrane filter and including one or more holes to allow the fluid sample access to said membrane filter; a flexible member (12) hermetically sealed to said support and spaced apart from said membrane filter to form a space between said membrane filter and said flexible member, said space being enclosed by said membrane filter (24), said support (22) and said flexible member (12), said flexible member being adapted to flex toward and away from said membrane filter; and an absorbent hydrophilic material (20) occupying a portion of said space and positioned to contact said membrane filter when said flexible member is flexed toward said membrane filter to promote passage of said fluid sample through said membrane filter.

Inventors: CLARK PHILLIP, CLARK, PHILLIP
Applicants: MILLIPORE CORP, MILLIPORE CORPORATION
Application year: 1991
Main classification: B01L3
IPC codes: G01N33, C12M1, B01D24