A DEVICE AND METHOD FOR SELF CONTAINED SOLID PHASE IMMUNODIFFUSION ASSAY

Abstract: A device and method for self containing solid phase immunodiffusion assay are comprised of a sample collector (15) and a pre-fabricated laminate (33) which can be used in many different forms. For example, the sample collector and laminate can be used with a tube having compartmentalized reagents. The seals can be broken through pressure on the sample collector. The sample collector is pushed through the seals, mixed with reagent, and then pushed into a ligand receptor reaction area which is part of the laminate. The tip of the sample collector contacts diffusable porous membranes or filters (25) and transfers the reactants to a capture membrane (18) wherein a ligand receptor reaction can be examined by the naked eye or otherwise.

Description: A DEVICE AND METHOD FOR SELF CONTAINED SOLID PHASE IMMUNODIFFUSION ASSAY CROSS-REFERENCE TO RELATED APPLICATION This is a continuation—in-part o-f United States Applicati Serial No. 938,003 filed December 3, 1986, the entire disclos of which is hereby incorporated by reference as if here set fo in full. BACKBROUND OF THE INVENTION In order to determine the condition of a patient, and minimize the diseased state, the need for a rapid diagnosis appropriate treatment by health care pro essionals is apparen Diagnosis of many conditions can be facilitated through t determination or quantitation of antibodies, antigens, nucleoti fragments, and other analytes from a biological specimen, whi are indicative of a particular disease state or condition. rapid, sensitive, specific, and simplistic assay is extreme useful for emergency situations, field testing, physicia offices and in home diagnostics. As diagnostic tests become mo simple and easier to perform, they are being performed away fr the professional clinical laboratory setting to physicia offices and even to the home, where untrained or poorly train individuals perform the tests usually following product inse instructions alone. These assays are useful provided they a performed properly and are safe to handle for the user. Assa that require multiple steps, have multiple reagents, and ha limited storage conditions are prone to misuse, especially they are performed by individuals without adequate training skills. Many types of ligand receptor assays have been developed a commercialized. These assays are less expensive if capit equipment can be eliminated, such as scintillation counter fluorometers, and colorimeters in the case of radiαimmunαassa fluorescent immunoassay, and enzyme immunoassay respectively. N instrumental assays, such as latex agglutination, enzy immunoassays on strips, tubes, membrane or filters have increas the usefulness and ease of performance of immunodl gnost testing, but are still cumbersome requiring washing step multiple reagent additions and usually refrigerated storag conditions. In some assays amplification or growth of viruses and bacteri are desirable before testing to increase the sensitivity o detection. In other assays adsorption steps to remov interferring substances or inhibitors of the ligand recepto assay, or long incubation of reagents are necessary to perform a assay. Each step for an assay increases the difficulty of testin for the minimally trained individual and any device that woul reduce user error would improve diagnostic testing. Horrisberger et al (J« Histo Cytochem volume 25: 295—305 1977) described the use of colloidal gold particles in a immunoassay. Leuvering in US patent number 4,313,734 als describe such an immunoassay. Cerny in PCT/US patent applicatio number 85/02534 describes a solid phase diffusion assay usin gold sol particles as an immunσlabel which can be visualized b the naked eye on a capture membrane, and requires no washin step. Bernstein et al (86th annual American Society fo Microbiology Meeting, 1986) presented and described a rapi immunσdi usion enzyme labeled antibody assay for GrOup streptococci on a membrane in which there is no washing step Gould and Zuk in US patent number 4,552,839 describe the use o colored or dyed beads in a solid phase immunoassay. Through th introduction of colored im uπαlabe led binding reagents (i.e gold sol particles, dyed particles, dye encapsulated liposomes etc.) and the removal of washing steps it becomes possible t perform receptor ligand assays in a closed system with th sequential additions of all reagents within that system. A number of antigens of interest in the diagnosis o infectious disease are collected with a sterile swab on a.shaf to remove the organisms from the suspected infected area or tes site (wounds, lesions, blood, tissues, pus, fluids, etc.). Th swab is generally used to transfer organisms to a suitable medi for culturing which may take as long as 4B hours for growth o bacteria, and 2 weeks for viruses. If the organisms are viabl and do grow, then their i entification could be made biochemical, morphological or immunological methods. This ti consuming method is slowly becoming replaced by more rap immunological testing methods or DNA probe methodologies. In many immunoassays that utilize a swab for collection antigens or cells, the swab is placed in a solution to relea the antigenic materials or cells after collection. It may necessary to use enzymes, acids, detergents, etc. to solubili or breakdown the antigens to expose antigenic determinants. T extracted material can then be used in an immunoassay by removi the fluid from the swab and mixing it with other reagents adding the other reagents directly to the swab extract. , In t case where membrane or filters are used to capture t immuπoreactants, it is necessary to bring the fluid containi the immunoreactants in contact with the filter or membrane. In addition, where extraneous cells or debris may interfer with an assay, it may be necessary to have a prefilter (large pare size filter or membrane) present between the swab and th capture membrane or capture filter to retain these unwante components. In some assays, where antigen expression may be low ampli i ation can be achieved if the organisms are first culture and then tested. If the culturing and the testing could b performed in a single device, then testing would be simplified In some assays where there are inhibitors, cross reactiv products, or clotting factors, red blood cells, etc., it may b necessary to add adsorbant materials (i.e. beads, kaolin antibody coated particles, antigen coated particles, or lecti coated particles) , anticoagulants, or buffers etc. before th ligand receptor assay can be performed. BRIEF DESCRIPTION OF THE INVENTION It is an object of the present invention, in one preferre form, to provide a novel test device that utilizes a swab o swab-like material (a shaft with a porous or fibrous absαrban material at one end) to collect a sample and to be able to react the swab with all the necessary reagents which are include within the device, and then to use the swab to transfer th reactants sequentially to other reactants if necessary, an finally to a reaction zone where the specific labelled reactaπ can be captured and visualized. It is another object of the present invention, in a secon preferred form, to provide a prefabricated assay device in th form of a laminate. This laminate in cooperation with a swab o other sample collector device provides a means for performin many types of ligand receptor assays. The laminate can be used i many different assay formats. It is a further object of the present invention to provide test device useful in performing ligand receptor assays tα detec antigens, haptεns, antibodies, DNA or RNA fragments, wherein th user is not required to dispense any of the reagents. It is a particular object of the present invention to provid a test device that can be stored at noπrefrigerated temperatures and can. be utilized to perform an assay on a biological specime or fluid without any additional reagents having to be provided t the test device. In addition it is a further object of the present invention t provide a test device which can utilize lyαphilized reagents tha can be reconstituted in situ within the device. The present invention maximizes the safety and ease o performance of ligand receptor assays through the use o apparatus designed to enable a biological specimen to be obtaine by a collection device comprising a shaft and an attached tip o adsorbent or absorbent porous or -fibrous material (i.e. rayon dacron, cotton swab). In the first form of the invention, the ti is inserted into a cylindrical tube. The cylindrical tub contains a sealed vessel, or chamber, or plurality of seale vessels or chambers in sequential order. The seal will break awa or collapse when pressure of the collection device (swab) i exerted on the seal by physically pushing the collection devic into and through each vessel. These sealed vessels may conta media, extraction reagents, diluents, labelled antibodie labelled antigens, labelled lectins, anticoagulants, adsαrbant inactivators, etc. which mix with the biological specim collected on the collection device. The reagents in these vesse may be lyophilized, enabling long term storage at n refrigerated temperature. The vessels are fixed in position the cylindrical tube to enable the seals to be broken wh physical pressure is exerted on the shaft of the collecti device. The collection device holder has appropriate stop poin to allow for the collection device tip to enter the appropira vessel and mix with its contents. A key feature of the vessel are that the tip and shaft of the collection device can pa through each of the vessels into a lower portion of t cylindrical tube and attached lower portion comprising a liga receptor reaction area. This ligand receptor area is part of pre—fabricated laminated assay device. The ligand receptor area is comprised of a capture membrane o a filter that will allow unbound reactants to pass through b diffusion and retain the appropriate labelled members of th binding pair. The capturing membrane or filter may be coated wit a member of the binding pair to capture the reactants. If captur particles are used, then the capture filter is utilized to retai the particles and allow unbound free labelled antigen or antibod to diffuse through. A prefilter may be used between the collecto tip and the capture or filter to remove any nonspecific bindin due to debris. An additional absσrbant material can be place behind the capture membane to increase the uptake of fluid. I either case a specific volume of reactants can be absorbed b controlling the ' size of the filter and absorbant materials. Th confi ura ion of the lower portion allows the collection devic to come into physical contact with the prefilter, captur membrane or capture filter. The pre—fabricated assay device i comprised of sequential layers of at least one porous membrane an impervious shield having at least one opening, and a filte means. DESCRIPTION OF THE DRAWINGS Fig. 1 is a cross—sectianal view of a first form of the invention showing the collection device holder, the collection device, th tube, the sealed reagent compartments and the lower ligan receptor transfer area; Fig. 2 is a perspective view of the basic structure of th collection device holder and collection device of Fig. 1 including the grooves far guiding the movement of the collectio device through the apparatus; Fig. 3 is a perspective view of the basic structure of the tub of Fig. 1, its compartmentalized reagents, and the nodule whic fits into the groove of the collection device holder; Fig. 4 is a perspective view of the sealed compartments (i.e. vessels) of the apparatus of Fig 1; Fig. 5 is a cross—sectional side view of the lower portion αf th apparatus of Fig. 1 showing the final position of the collection device tip at the window of the ligand receptor area; Fig. a is an exploded perspective view αf the ligand recepto test area as used in the apparatus in Fig 1; Fig. 7 is an exploded perspective view of the pre— abricate assay device according to a second form αf the invention; Fig. 8 is a pictoral view αf the sample collection -device i contact with the pre-fabricated assay device using the secon form of the invention; Fig. 9 is a perspective view of a prefabricated assay device an sample collection device being sandwiched between an uppe hydrαphobic pressure plate 29 and a lower flat hydrophobic plat 30 according to a third form of the invention; and Fig. 10 is a perspective view of a prefabricated assay devic showing a modification of the third form of the invention havin a sample collection device positioned on a lower flat hydrophobi plate 30 which is hinged to a hydrophobic upper pressure plat 29. DESCRIPTION OF THE PREFERRED EMBODIMENTS In the following description, the various forms of th apparatus and the method of the invention will be described i exemplary terms only, for an antigen determining an immunoassa test. This discussion, however, is simply to illustrate th structure and use of the apparatus and the technique and steps o the method. The various forms of the apparatus clearly can b used for any ligand receptor assay in which washing steps hav been eliminated and transfer of the reactants to or through porous membrane or filter is used. The best modes, as describe hereinafter, are accordingly, to be considered exemplary and no limiting as to the scope and concept of the invention. Referring first to Fig. 1 for a general depiction of th apparatus, the inventive apparatus comprises a collection devic holder 14 which is comprised of a restrictive portion 1 that hol the shaft of the collection device 2 in place, a cylindrical tub 13 which is comprised of one or more sealed reagent compartment 15 and 20, and a lower ligand receptor reaction area 10. Area 1 is drawn enlarged, but can be smaller, see Fig. 5 for example. Referring to Fig. 2, the collection device holder 14 has nodule 16 which positions onto the cylindrical tube and prevent the apparatus from being accidentally opened. Nodule 16 can b heat sealed or made to cooperate with a .groove formed in tube 13 or the cooperating nαdule 17 shown in Fig. 3. or other means ca be provided to accomplish the sealing function. When a saπtple i to be taken, the collection device holder is removed an separated from the cylindrical tube by twisting and pulling up o the collection device holder. This frees up the collection devic holder which is then used to collect the test sample (i.e. throat swab, pus, blood, urethral swab, etc.) by allowing th collection device tip 5 to come into contact with the suspec tissue, fluid, wound, etc. Referring to Fig. 2 and Fig. 3, after obtaining a test sample, the collection device holder is replaced onto the cylindrical tube 13 and turned until the nodule 4 (Fig. 3) on the cylindrical tube is in alignment with the groove 18. The collection devic holder is then manually forced downward until the nodule 4 stop at the horizontal groove 19. When the nodule 4 is in contact wit horizontal groove 19, then si ultaneously the tip 5 will hav broken through the first seal (Fig. 4) , mixing with the content of the first chamber 15, then breaking through seal 7 an emptying its contents into chamber 20. The number of independen chambers is related to the number of required reagent addition and incubation steps. One chamber or a plurality αf chamber could be used and the mixing of reagents controlled using th principles of nodule 4 and grooves 18, 19 as previously descibed. In the preferred embodidment, the collection device holder i turned to the right and then back and forth using groove 19 t mix the contents of vessel 20 through the simultaneous turning o the collection device tip. Referring to Figs. 2 and 3, after an appropriate incubatio time, the collection device holder 14 is turned to the right an thus aligning nodule 4 (Fig. 3) with groove 3 (Fig. 2) and the manually farced downward until the movement of nodule 4 i I stopped by the groove end 21 (Fig. 2) . Referring to Figs. 5 and 6 the lower portion 10 may b physically one piece with the cylindrical tube 13 or may be a attached separate piece. When the nodule 4 is in contact with th groove end 21, then the collection device tip 5 is in contac with the prefilter membrane 25 through . the window 11. Th reactants flow through the prefilter membrane through holes 24 o- impervious shield 23 which holds the prefilter membrane 2 against hole or window 11. The shape of the lower portion 10 i configured to enhance contact of the collection device tip 5 wit the prefilter or reaction membranes. If preferred, the prefilte could be placed an the inside wall αf the window 11. In any case, the reactants flow through holes 21 and 22 αf shield 20 whic holds membranes 18 and 19 respectively in place. The holes 21 an 22 restrict the flow of the reactants through a capture membran 19 and a control membrane 18 and enhance the signal iof t reaction by concentrating the labelled ligand or receptor bindi pairs into a small area. Absorbant 17 absorbs excess flu diffusing through the membranes. When an appropriate volume fluid has diffused through the membranes, usually by saturati of the absorbant, the capture and control membrane are examin within the holes 21 and 22 respectively by lifting the tab 28 the adhesive tape 12. In the preferred form αf the invention, the assay results a examined by the naked eye since color changes are the resul However the invention is not limited in this manner and t "examination" of test results could be done with instruments otherwise dependent upon the protocol of the particular assay. Adhesive tape 12 holds the absorbant 17 in place and appli the necessary pressure tα insure diffusion of fluid through t various layers of the ligand receptor test device 33A. The col intensity of the capture membrane IB is compared tα the col intensity of the control membrane 19. A positive result determined by viewing a more intense color in the captu membrane than in the control membrane. A negative result determined by no significant color or the same weak color in t capture and control membranes. In competitive inhibition assa the positive and negative results are reversed. In t performance of drug analyte assays, the size of the ring of col in a single larger capture membrane is related to t concentration of drug in the test sample. The design of t ligand receptor area , the coating of reagents on the membrane and the addition or deletion of capture or control membranes a dependent on the particular type of assay being performed. T capture membrane can be coated with antigen or antibody, or oth complementary ligands or receptors and can be used tα determin the presence of different antigens or antibodies. The number chambers used in the apparatus is dependent upon the type o assay and can contain diluents, media for growth amplification o mi roorganisms, lyophilized labelled ligands or receptors, etc The seal 7 (Fig. 4) may be attached to two chamber simultaneously or may be independent. Therefore the chamber cαuld be attached to each other or be independent. Referring tα Fig. 6, the prefilter 25 is -fixed around th periphery of hole 11 in tube lO by taping, gluing, heat sealing, chemically sealing, sonic welding, or the like. Impervious shiel 23 is a single side adhesive coated laminate having adhesive o the side facing the prefilter 25. The impervious shield laminat 23 with hole 24 fixes and exposes the prefilter to the tube 1 and hale 11. The impervious shield 20 can also be a single sid adhesive coated laminate which fixes membranes IB and 19 ont absorbent 17. Absorbent 17 is fixed on the side opposite to th positioned membranes with adhesive tape 12. The adhesive tape 1 fixes the assay device 33A to tube 10. Alternative m^aπs o fixing can include clamps, glues, or the like. In order t examine a test result on the membranes 18 and 19 it is necessar to separate the impervious shields 23 and.20 from each other t expose the membranes. By pulling back adhesive tape 12, a portio of the assay device 33A consisting of shield 20, membranes 18 an 19, and absorbent 17 separates from filter 25 and shield 23 thereby exposing membranes 18 and 19 for examination. Referring to Fig. 7, the pre-fabricated assay device 33 i a laminate comprised of a prefilter membrane 25 held in place b a hydrophobic impervious shield 23 having a hale 24 through whic reactants from absorbent tip 5 can flow into the prefilter. Whe absorbent tip 5 is pressed into contact with prefilter 25, th flexible prefilter is forced into contact through hale selectively clustered and sized 21 and 22 αf impervious shield 2 with control and capture membranes IB and 19. The fluid reactant from tip 5 flow from prefilter 25 to bath membranes IB and 1 simultaneously. Absorbent 17 is in contact with membranes IB an 19 and the fluid reactants diffuse through membranes IB and 19 t absorbent 17. At the conclusion αf a ligand receptor assay th impervious shield 23 with attached prefilter 25 is removed an membranes 18 and 19 are examined by the naked eye, or otherwise, for any color change, or the like. Fig. 8 is a perspective view of a collection device ti being manually brought into contact with an assembled p fabricated assay device 33. The fluid of a labelled ligand receptor reagent in combination with sample is transferred f tip 5 through prefilter membrane 25 and through control capture membranes IB and 19 to absorbent 17. This shows general utility of the invention devices 33 and 33A to be used many different environments. Fig. 9 is a perspective view of a sandwich device 36 holding collection device tip 5 under pressure against a p fabricated assay device 33. Hydrophobic upper pressure plate is movable and hinged to hydrophobic lower flat plate 30. Af reacting sample collection device tip 5 with labelled ligand labelled receptor by manipulating shaft 2, the sample collect device tip 5 is positioned horizontally in contact with prefil membrane 25 and pressure is applied to the device forcing absorbent tip 5 to push the prefilter 25 into contact with pύ membranes 18 and 19 and allowing the reactants to flow thro and into the pre-fabricated assay device 33. Pressureιcan applied to pressure plate 29 by means of a weight, cla manually squeezing, or the like. After an appropriate period time to allow the fluid reactants to penetrate the assay devi 33, pressure is removed from the hinged sandwich device and t impervious shield 23 is lifted to allow the control and captu membranes of the pre—fabricated assay device to be examined. Fig. 10 is a perspective view of a hinged sandwich device for applying pressure to sample collection device tip 5 and t pre-fabricated assay device 33. Upper hydrophobic pressure pla 29 is contoured as at 31 to the sample collection device 2 ,and to maximize the amount of fluid that can flow into the pr fabricated assay device 33 which is positioned on the low hydrophobic plate 30. After the sample collection device h reacted with labelled ligand or labelled receptor the samp collection device is manually placed within the hinged sandwi device 37 with sample collection device tip 5 in contact wit prefilter membrane 25 and pressure is applied tα and maintaine by the hinged sandwich device 33. The hinged sandwich device 3 consists of a hinge 32 and a clamping mechanism such a protrusions 34 from upper pressure plate 29 and a complementar fitted piece such as the orifices 35 which will produce friction fit and maintain the closed clamped position of th hinged sandwich device 33. After an appropriate time period th hinged sandwich device is opened and the prefilter membrane 25 i removed tα expose the control and capture membranes of the pre fabricated assay device 33. In Figs. 9 and 10, the assay device 33 can be layed loosely or affixed as by glueing or the like tα one of the plates 29 o 30. Also a farm fitted depression to hold the device 33 could b formed in a plate 29 or 30. DEFINITIONS In the following section certain terms used in th speci-fication and claims herein shall be understood to have th following meanings: Ligand: Any organic compound for which a receptor naturall exists or ' can be prepared including haptens, antigens sugars, vitamins, pεptides and the like. Receptor: Any compound capable of recognizing a specifi molecular con iguration of a ligand including antibodies lectins, enzymes, nucleic acids, Fab fragments, avidin an the like. Ligand receptor reaction: Any binding between a receptor ≤lnd it complementary ligand. Porous membrane: any porous solid matrix including porous spreading layers, bibulous papers, filters, and th like. Coated porous membrane: A porous membrane that has a ligand o receptor covalently ar nαncovaleπtly attached to it surface. Impervious shield: any hydrophobic material that will not all diffusion of fluid to occur through its surface. Thi material can be plastic, a plastic adhesive laminate, an the like. Chamber: a partitioned space, vessel, reservoir, compartment o the like. The following example is illustrative: EXAMPLE 1 A RAPID I MUNODI GNOSTIC TEST FOR GROUP A STREPTOCOCCI. Group phage associated lysin enzyme which is effective in fragmentin and solubilizing the Group A streptocσccal pαlysaccharide wa diluted in a buffer of .05M citrate phosphate pH 6.1 containin 0.005 EDTA, 0.005 DTT, 0.17. rabbit IgG, 0.05% sodium azide an mixed with rabbit anti streptαcoccal Group A coated gold so particles (0D aiβ 1.5) diluted in a buffer of 0.02 Tris pH B. containing 1.07. BSA, 0.27. sodium heparin, 0.37. n acetylglucosamine and 0.02.7. sodium azide in a ratio of 3 part lysin reagent to 1 part antibody gold sol reagent. The combine reagent was sterile filtered through a 0.2 micron cellulos acetate filter and 200 microuters were aliquoted into acryli walled reaction cup vessels, having an aluminum foil seale bottom. The aliquots were frozen and lyophilized. The reactio cup vessels were sealed with aluminum foil and contact cemen under nitrogen. Another reaction vessel was cemented to th aluminum fail lid of the first vessel. Two hundred microliters o distilled water was added to the second vessel and then cemente and sealed with aluminum foil. The vessels were placed an positioned into the cylindrical tube. The ligand receptor are was prepared by coating nitrocellulose membranes with rabbit ant Group A streptococcal antibody for the capture membranes, an normal rabbit immunαglobulin for the control membrane. Th membranes were dried and fixed to a- diacetate laminate which ha 1.5mm diameter holes for each membrane. A 1.2 micron cellulos acetate prefilter was used tα cover the hole (window) in th lower portion of the tube. A dacrαn tipped swab was seeded wit varying conceπtrations αf group A streptococci. The swab wa placed into the cylindrical tube and forced downward to break th first two seals an the reaction vessels. The swab was incubate for 4 minutes at room temperature allowing the lysin enzyme t solubilize the Group A streptococcal polysaccharide and th reaction αf the gold labelled anti Group A antibody to far complexes with the released polysaccharide. After the fou " minutes, the swab was farced downward through the third seal int the lower portion, coming in contact with the ligand recepto area 33. The fluid diffused through the prefilter into th capture and control membranes. After 30 seconds the tab 28 on th ligand receptor area 33 was pulled away from the lower portio and examined by eye. A distinct color reaction with 2 κ 10 organisms of Grαup A streptococci could be distinguished in th capture membrane compared tα the colorless control membrane. The foregoing disclosure and the showing made in the drawing are merely illustrative αf the principles αf this invention an are not tα be interpreted in a limiting sense. It is understoo that through the example and embodiments described herein, tha various modifications in light thereof will be suggested t persons skilled in the art to be included in the spirit an review of this application and the scope of the approved claims.

Claims: AMENDED CLAIMS [received by the International Bureau on 06 May 1988 (06.05.88); original claim 2 cancelled; claims 1,3,5-6,10,12 and 13 amended; other claims unchanged (7 pages) ] 1. (Amended) A self-contained ligand receptor assay, the combination comprising; a) a collection device having an elongated shaft with an absorbent tip for collecting a specimen; b) a tube, cooperative with said collection device, said tube having an interior, an open end and a distal endr c) at least one sealed chamber in said tube; d) a number of reagents equal to the number of said at least one chamber located one reagent in each said at least one chamber; e) means to seal each said reagent in a respective one of said at least one chamber; f) said seal means comprising frangible seals; g) ligand receptor reaction means including at least one porous membrane; h) a hole formed in said tube in predetermined spaced relation to said distal end thereof; i) means to fix said ligand receptor reaction means to said tube covering said hole with said least one porous membrane exposed to said interior of said tube; and j) the length of said shaft being such as to permit said absorbent tip to reach through all of said at least one chamber to mix all of said at least one reagent and to permit fluid diffusion between said tip and said at least one membrane; whereby ) a labelled ligand or a labelled receptor is immobilized on said ligand receptor reaction means and th result of said assay can be examined. 2. (Cancelled) 3. (Amended) The device of claim 1, a guiding mechanis for said collection device, said guiding mechanism comprising guide and stop means, nodule means on said tube, and said guid and stop means and said nodule means cooperating with each othe causing sequential breaking of said at least one frangible sea and moving said absorbent tip completely through at least one chamber when said absorbent tip passes through all of said at least one chamber. 4. The device of claim 1, wherein said at least one reage comprises a labelled ligand or a labelled receptor. 5. (Amended) The device of claim 4, further comprising a least one other chamber, and microbiological growth media in sai other chamber. 6. (Amended) The device of claim 4, further comprising a least one other chamber, and an extraction reagent for exposing antigenic determinants or hidden epitopes of an antigen in said other chamber. 7. The device of claim 4, wherein said labelled ligand o labelled receptor reagent is lyophilized. 8. The device of claim 4, wherein said labelled ligand o labelled receptor reagent comprises a chromophore. 9. The device of claim 8, wherein said chromophore is selected from the group consisting of dyes, dyed particles, pigments, metal sol particles, or dye encapsulated liposomes. 10. (Amended) The device of claim 9, wherein said chromophore is a metal sol particle selected from the group consisting of gold, silver, or the combination of gold and silver. 11. The device of claim 1, wherein said absorbent tip for collecting a specimen is sterile. 12. (Amended) The device of claim 4, further comprising a least one other chamber, and adsorption particles in said other chamber, said adsorption particles comprising binders to remove inhibiting or cross reactive substances from said assay; and at least one other porous membrane, and a) said adsorption particles are being larger than said labelled ligands or labelled receptors, and b) said at least one other porous membrane having a effective pore size smaller than said adsorption particles and larger than said labelled ligands or labelled receptors. 13. (Amended) The device of claim 1, wherein said at leas one porous membrane is a plurality of porous membranes at least one membrane of which is coated with different ligands or receptors. 14. The device of claim 4, wherein one of said at least on chamber contains ligand or receptor coated particles which are larger in mean particle size diameter than said labelled ligand labelled receptor and the effective pore size of said at least o porous membrane. 15. A method of performing a ligand receptor assay using sample collector means, comprising the steps of; a) putting a specimen on said sample collector means; b) contacting said specimen on said sample collecto means with one or more reactants as required to perform sai assay; c) providing at least one porous membrane as require to perform said assay; d) causing liquid diffusion from the reactants on sai sample collector means after said step b) to all of said at least one porous membrane simultaneously; and e) examining all of said at least one porous ntømbran after performance of all of steps a) to ' d) above; whereb an color change or the like caused by a labelled ligand or a labelled receptor being captured or not being captured on said at least one membrane in accordance with the protocol o said assay can be readily observed. 16. The method of claim 15, and the additional steps of providing two of said porous membranes, and causing one of said two porous membranes to be a control membrane and the other of said two porous membranes to be a capture membrane. 17. The method of claim 15, and the additional step of filtering said reactants during said step d) using filter means having a pore size selected to be large enough to permit said liquid diffusion of said step d) and small enough to prevent debris larger than said selected pore size and the like which ma be on said sample collector means from contacting any of said at least one porous membrane. 18. The method of claim 15, and performing said step d) including the sub-step of creating physical pressure at least indirectly between said sample collector means and all of said at least one porous membrane simultaneously. 19. The method of claim 15, wherein said steps c) and d) ar performed using a pre-fabricated assay device, and the steps of constructing said assay device in the form of a laminate comprising sequential layers as follows: i) said at least one porous membrane; ii) impervious shield means, and the steps of providin at least one opening in said impervious shield means equal i number to the number of said at least one porous membrane, and sizing and clustering said at least opening so that said step d) can be performed through all of said at least one opening simultaneously; and iii) filter means, and selecting a pore size for said filter means large enough to permit liquid diffusion of jsai step d) and small enough to prevent debris larger than said selected pore size and the like which may be on the sample collector means from contacting any of said at least one porous membrane. 20. The method of claim 19, and the additional steps performed during said construction of said laminated device step of providing an absorbent means layer, and locating said absorben means layer next to said at least one porous membrane on the side thereof opposite said impervious shield means. 21. A pre-fabricated assay device in the form of a lamina comprising sequential layers of: a) at least one porous membrane; i) impervious shield means formed with at least one opening equal in number to the number of said at least one porous membrane; ii) means for sizing and clustering said at least one opening so that liquid diffusion between said at least one porous membrane and a reactant brought into contact with said assay device can occur through all o said at least one opening simultaneously; and c) filter means, and said filter means having a por size large enough to permit said liquid diffusion and smal enough to prevent debris larger than said selected .poip. siz from contacting any of said at least one porous memb_.aj_ . 22. The device of claim 21, and a sample collection device comprising an elongated shaft and an absorbent tip. 23. The device of claim 22, and a container comprised of liquid labelled receptor or labelled ligand reagent. 24. The device of claim 23, wherein the said labelled reagent comprises a chromophore. 25. The device of claim 24, wherein the said chromophore i selected from the group consisting of dyes, dyed particles, pigments, metal sol particles, or dye encapsulated liposomes. 26. The device of claim 22, and a sandwich device, said sandwich device comprising a pair of plate means, means to hinge said plate means together along one respective side of said pair of plate means, positioning said pre-fabricated assay device between said plate means, and means to apply pressure to said ti when said tip is inserted between said pair of plate means and i contact with said pre-fabricated assay device. 27. The device of claim 21, and said laminated assay devic including an absorbent layer in contact with said at least one porous membrane on the side thereof opposite said impervious shield. 28. The device of claim 26, and said laminated assay devic including an absorbent layer in contact with said at least one porous membrane on the side thereof opposite said impervious shield. 29. The device of claim 26, said means to apply pressure comprising clamping means.

Inventors: BERNSTEIN DAVID, BERNSTEIN, DAVID
Applicants: NEW HORIZONS DIAGNOST CORP
Application year: 1987
Main classification: G01N33
IPC codes: A61B10, A61F13, C12M1, G01N33