Claims for Patent: 5,217,905

Summary for Patent: 5,217,905

Title:	Device and method for the rapid qualitative and quantitative determination of the presence of a reactive ligand in a fluid
Abstract:	A device for the rapid qualitative and quantitative determination of the presence of a reactive ligand in a fluid. This device comprises a first reaction zone in which there is an at least temporarily impermeable membrane designed to receive a sample of test fluid and to be associated with at least one labeled reagent; a second reaction zone which is bounded on the one hand by the said membrane and on the other by a second at least temporarily impermeable membrane comprising a solid phase containing a reference reagent; and a third reaction zone which contains means for developing the reaction. A method for the rapid qualitative and quantitative determination of the presence of a reactive ligand in a fluid. Applications to the detection of the presence, in a biological fluid, of antibodies or antigens in particular.
Inventor(s):	Marchand; Joseph (Verrieres Le Buisson, FR), Toledano; Jacques (Paris, FR)
Assignee:	Compagnie Oris Industrie S.A. (Gif-Sur-Yvette, FR) Cistest (Paris, FR)
Application Number:	07/220,895
Patent Claims:	1. A device for rapid detection of the presence of a reactive ligand in a fluid test sample, comprising: a first reaction zone for receiving the fluid test sample and containing a temporarily impermeable first membrane, a labeled reagent capable of binding to the reactive ligand to be detected, and permeating means for rendering said first membrane permeable; a second zone, separate from said first reaction zone and temporarily isolated by said first membrane, said second zone containing a solid phase having bound thereto an unlabeled reference reagent capable of recognizing and retaining labeled reagent from said first zone which has not bound to the reactive ligand to be detected, and a second temporarily impermeable membrane opposite from said first membrane; and a third reaction zone, separate from said second reaction zone and at least temporarily isolated therefrom by said second membrane, said third reaction zone including disclosing means for disclosing the presence or absence of said labeled reagent; wherein said permeating means comprises an enzyme capable of dissolving of lysing said first and second membranes, said enzyme of said permeating means being different from the label of said labeled reagent, and wherein said first and second membranes are made of a material which is impermeable for a period of time sufficient to enable a ligand-labeled reagent or labeled reagent-reference reagent reaction to occur and is subsequently permeable due to said permeating means to allow fluid to pass therethrough to a subsequent reaction zone. 2. The device according to claim 1 wherein said permeating means is incorporated in said first membrane. 3. The device according to claim 1 wherein said labeled reagent is in solid form within said first reaction zone and is supported by said first membrane. 4. The device according to claim 1 wherein said solid phase in said second reaction zone consists of an insoluble support to which said unlabeled reference reagent is bound and said unlabeled reference reagent corresponds to the ligand to be detected. 5. The device according to claim 1 wherein said labeled reagent is labeled with a material capable of reacting with a chromogenic substance to produce color, and wherein said disclosing means comprises a chromogenic substance which reacts with said labeled reagent when present in said third reaction zone to form a chromogen which can be read by colorimetry. 6. The device according to claim 1 wherein said disclosing means includes quantifying means for quantifying the presence of said labeled reagent in the third zone. 7. The device of claim 6 wherein said quantifying means is a spectrophotometer or a radioimmunoassay quantifying apparatus. 8. The device according to claim 1 wherein said reaction zones are superimposed to form an upper stage comprising said first reaction zone, a middle stage comprising said second reaction zone, and a lower stage comprising said third reaction zone. 9. A method for rapid detection of the presence of a reactive ligand in a fluid by means of the device of claim 1, which comprises the steps of: adding a sample of fluid for analysis to said first reaction zone and maintaining said sample in said first zone for a sufficient time to enable a ligand-labeled reagent reaction to occur between said labeled reagent and any reactive ligand in said sample; causing said first membrane to become permeable to the fluid after said ligand-reagent reaction has occurred by means of said permeating means, to thereby permit transfer of said fluid from said first reaction zone to said second zone; maintaining said fluid in said second reaction zone for a sufficient time to enable a labeled reagent-reference reagent reaction to occur between any of said unbound labeled reagent in said fluid and said unlabeled reference bound to the solid phase in said second zone; causing said second membrane to become permeable to the fluid, after the labeled reagent reference reagent reaction has occurred, by means of said permeating means, to thereby permit transfer of said fluid from said second reaction zone to said third reaction zone, said solid phase being retained in said second zone; and detecting the presence of said labeled reagent in said third zone by means of said disclosing means, wherein the presence of said labeled reagent indicates the presence of said reactive ligand. 10. A method in accordance with claim 9 wherein said first and second membranes are made of gelatin and said permeating means comprises collagenase. 11. The device according to claim 1, wherein said permeating means is in said first reaction zone, and not incorporated in said first membrane. 12. A device in accordance with claim 1, wherein said labeled reagent is labeled with a label which can be read by fluorescence, chemiluminescence or radioactivity. 13. A device in accordance with claim 1 wherein said first and second membranes are made of gelatin and said permeating means comprises collagenase. 14. A device for detecting the presence of a reactive ligand in a fluid test sample at a semi-quantitative level, comprising: a first reaction zone for receiving the fluid test sample and containing a temporarily impermeable first membrane, a predetermined quantity of unlabeled reagent capable of binding to the reactive ligand to be detected, and permeating means for rendering said first membrane permeable; an intermediate reaction zone containing a labeled reagent capable of binding to the reactive ligand to be detected, said intermediate zone being separated from said first reaction zone by said first membrane to prevent contact of the sample in said first reaction zone with said labeled reagent for a time sufficient to enable a ligand-unlabeled reagent reaction to occur; a second reaction zone, separate from said first and intermediate reaction zones, said second zone containing a solid phase having bound thereto an unlabeled reference reagent capable of recognizing and retaining labeled reagent from said intermediate zone which has not bound to the reactive ligand to be detected, and a second temporarily impermeable membrane; and a third reaction zone, separate from said second reaction zone and at least temporarily isolated therefrom by said second membrane, said third reaction zone including disclosing means for disclosing the presence or absence of said labeled reagent; wherein said permeating means comprises an enzyme capable of dissolving or lysing said first and second membranes, said enzyme of said permeating means being different from the label of said labeled reagent, and wherein said first and second membranes are made of a material which is impermeable for a first period of time sufficiently to enable a ligand-unlabeled reagent or labeled reagent - reference reagent reaction to occur and is subsequently permeable due to said permeating means to allow fluid to pass therethrough to a subsequent reaction zone, whereby detection of the presence of labeled reagent by said disclosing means indicates the presence of the reactive ligand in the test sample in a quantity greater than the known quantity of unlabeled reagent in said first reaction zone. 15. The device according to claim 14 wherein said intermediate zone comprises at least one layer of vesicular structures having at least one labeled reagent which is capable of being released by said permeating means. 16. The device according to claim 14 wherein said intermediate zone is formed by an intermediate membrane intercalated between said first membrane and said second reaction zone, said labeled reagent being present between said first and intermediate membranes. 17. A device according to claim 14 wherein said reaction zones are superimposed to form an upper stage containing said first reaction zone, an intermediate stage below said upper stage, a middle stage below said intermediate stage, comprising said second reaction zone, and a lower stage comprising said third reaction zone. 18. A device in accordance with claim 14 wherein said first and second membranes are made of gelatin and said permeating means comprises collagenase. 19. A method for rapid detection of the presence of a reactive ligand in a fluid at a semi-quantitative level, by means of the device of claim 14, which comprises the steps of: adding a sample of fluid for analysis to said first reaction zone and maintaining said sample in said first reaction zone for a sufficient time to enable a ligand-unlabeled reagent to occur between said predetermined quantity of unlabeled reagent and any reactive ligand in said sample; subsequently causing said fluid to contact said labeled reagent in said intermediate reaction zone after said ligand-unlabeled reagent reaction has occurred in said first reaction zone to enable a ligand-labeled reagent reaction to occur between said labeled reagent and any reactive ligand remaining in said samples; after said ligand-labeled reagent reaction has occurred, transferring said fluid from said intermediate reaction zone to said second reaction zone; maintaining said fluid in said second reaction zone for a sufficient time to enable a labeled reagent-reference reagent reaction to occur between any of said unbound labeled reagent in said fluid and said unlabeled reference reagent bound to the solid phase in said second zone; causing said second membrane to become permeable to the fluid after said labeled reagent reference reagent reaction has occurred, by means for said permeating means, to thereby permit transfer of said fluid from said second reaction zone to said third reaction zone, said solid phase being retained in said second zone; and detecting the presence of said labeled reagent in said third zone by means of said disclosing means, whereby detecting of the presence of labeled reagent in said third zone indicates the presence of the reactive ligand in the test sample in a quantity greater than the known quantity of unlabeled reagent in said first reaction zone. 20. A method in accordance with claim 19 wherein said first and second membranes are made of gelatin and said permeating means comprises collagenase.

Details for Patent 5,217,905

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Smith & Nephew, Inc.	SANTYL	collagenase	Ointment	101995	06/04/1965		2010-06-08