Patent EA-000409-B1 (EA)
Title: Three-dimensional colorimetric assay assemblies
Inventor: CHARYCH DEBORAH
IPC: G01N33/58
Filed: 1996-02-13
Abstract: 1. Three-dimensional polymeric multilayer assay assemblies, which change color in the presence of analyte comprising: a) ligands having direct affinity for the analyte or which can function as competitive binders to the analyte, b) linear structural linkers having two terminal ends, wherein said linkers are attached at their first terminal ends to said ligand moieties, c) a conjugated polymer backbone to which said structural linkers are bound at their second terminal ends, and d) ordering head groups which are bound to the surface of the conjugated polymer backbone in positions not occupied by the structural linker, wherein said assemblies are in the form of a liposome, a double-chain, braided, lamellar, helical, tubular, or fiber-like shape. 2. The assemblies of Claim 1, wherein the ligands having affinity for the analyses are selected from the group, comprising biomedical materials, pathogens, drugs, radioactive metals, or industrial materials. 3. The assemblies of Claim 3, wherein said biomedical materials are selected from the group comprising pathogens and cells infected by them, drugs, hormones, blood components, disease indicators, cell components, antibodies, lectins, enzymes, genetic material, and their metabolic derivatives. 4. The assemblies of Claim 3, wherein said pathogens are selected from the group comprising viruses, bacteria, and parasites. 5. The assemblies of Claim 4, wherein said viruses are selected from the group comprising influenza, cold, rubella, chicken pox, hepatitis A&B, herpes simplex, polio, small pox, plague, HIV, vaccinia, rabies, Epstein Barr, reovirus, rhinovirus, and mutations, liqand recognizable parts thereof. 6. The assemblies of Claim 5, wherein said bacteria are selected from the group comprising E. coli, tuberculosis, salmonella, streptococcus, and mutations, strains and degraded parts thereof. 7. The assemblies of Claim 5, wherein said parasites and other pathogens are selected from the group comprising malaria, sleeping sickness, river blindness, and toxoplasmosis. 8. The assemblies of Claim 1, wherein said ligand is provided for the detection of a pathogen analyte. 9. The assemblies of Claim 9, wherein the analyte is a virus. 10. The assemblies of Claim 9, where said ligand is selected from the group comprising, epidermal growth factor for vaccinia analyte, acetylcholine receptor for rabies analyte, complement receptor for Epstein Barr analyte, beta-adrenergic receptor for reovirus analyte, ICAM-1 for Rhinovirus analyte, polio virus receptor for polio virus analyte, trisaccharide analyte for cholera toxin analyte, tetrasaccharide for neutrophil analyte, and derivatives and analogues thereof capable of associating with an analyte. 11. The assemblies of Claim 8, wherein said ligand is sialic acid and its derivatives and analogs which will bind to colronaviruses, influenza virus, encephalomyelitis, chlamydia, sendi virus, mumps, Newcastle disease, myxovirus, encephalo-mycarditis virus, meningitis, or malaria. 12. The assay of Claim 8, wherein said said ligand: is selected from the group comprising tetrasaccharides, cell adhesion peptides, trisaccharides and transmembrane receptors. 13. The assemblies of Claim 8, wherein the ligand provided to detect HIV analytes is selected from the group comprising CD4, sCD4, CD26, vasoactive intestinal peptide, peptide T, sialic acid, and derivatives and analogues thereof capable of associating with HIV. 14. The assemblies of Claim 1, wherein said polymer is comprised of polymerizable lipid monomers. 15. The assemblies of Claim 14, wherein said monomers are chosen from the group comprising acetylenes, diacetylenes, alkenes, thiophenes, imides, acrylamides, methacrylates, vinylether, malic anhydride, urethanes, allylamines, siloxanes anilines, pyrroles and vinylpyridinium. 16. The assemblies of Claim 15, wherein said polymer backbone is comprised of diacetylene monomers. 17. The assemblies of Claim 1, wherein said ordering head groups are hydrophilic. with the capacity to mutually hydrogen bond. 18. The assemblies of Claim 18, wherein said ordering head groups are selected from the group comprising; -CH2OH, - CH2OCONHPh, -CH2OCONHEt, -CH2CH (Et) OCONHPh, - (CH2)9OH, - CH2OCOPh. -CH2 OCONHMe, -CH2OTs, -CH(OH) Me, -CH2OCOR2, wherein R2 is n-C5 H11, n-C7H15, n-C9H19, n-C11H23, n-C13H27, n-C15H31, n-C17H35, Ph, PhO, or o-(HO2C)C6H4, -OSO2R2, wherein R2 is Ph, p-MeC6H4, p-FC6H4, p-ClC6H4, p-BrC6H4, p-MeOC6H4, m-CF3C6H4, 2-C10H7, or Me -CO2M, wherein M is K, HNa or Ba/2, -CH2OCONHR2 or - CH2CONHR2, wherein R2 is Et, n-Bu, n-C6H13, n-C8H17, n-C12H25, cyclo C6H11, Ph, p-MeC6H4, m-MeC6H4, o-Cl6H4, m-ClC6H4, p-ClC6H4, o-MeOC6H4, 3-Thienyl, Me, Et, Ph, 1-C10H7, Et, Ph, EtOCOCH2, BuOCOCH2, Me, Et, i-Pr, n-C6H13, Et OCOCH2, BuOCOCH2, Ph, 2,4 (NO2)2C6H3OCH2 or CH2CH2OH. 19. The assemblies of Claim 17, wherein said ordering head groups are are carboxylic acid. 20. The assemblies of Claim 1, wherein the non-binding terminal of the monomers is selected from the group comprising: CH3-, CH3O, neo-C5H11O-, Cyclo-C6H11O, PhCH2O-, p-AcC6H4O-, p-BZC6H4O-, p-BrC6H4COCH2O-, p-(PhCH=CHCO)C6H4O-, p-(PhCOH=CH)C6H4O-, oBzC6H4NH-, p-BzC6H4NH-, MeOCH2CH2NH-, n-C6H13NH- and EtO-. 21. The assemblies of Claim 20, wherein the terminal is a methyl group. 22. The assemblies of Claim 1, wherein the assemblies are bound to the backbone due to associating the chemical groups of said assemblies with the backbone surface. 23. The assemblies of Claim 22, wherein the backbone is selected from the group: syphedex, silica gel, sepheros, polyacrynitriles, filters, gold, silicon chips, silica gel. 24. A method of making the three dimensional polymeric multilayer assay assemblies of Claim 1, comprising: a) combining diyne monomers with ligands in an organic solvent, b) evaporating the solvent, forming dry lipid group, c) repeated obtaining of suspension of said dry product in liquid selected from the group, comprising water and aqueous solution, forming reaction solution, d) heating the solution above the main-phase transition temperature of the diyne monomers, e) agitating the solution and cooling it to at least 4 degree C, f) polimerizing the reaction solution. 25. The method of Claim 24, wherein in step a), the solvent is selected from chloroform, benzene, alcohol, cyclohexane, methylene chloride, acetonitrile, and carbontetrachloride. 26. The method of Claim 24, wherein the aqueous solution of step c) is selected from water, buffer solution, cell media, physiological saline, phosphate buffered saline, Trizma buffer, HEPES, and MOPS. 27. The method of Claim 24, wherein before the cooling in step e), the solution is filtered. 28. The method of Claim 24, wherein the diyne-ligand mixture is cooled at between 4 degree  and -20 degree C for between 5 minutes and 5 hours. 29. The method of Claim 28, wherein the mixture is cooled at between 0 degree  and -15 degree C for between 5 and 20 minutes. 30. The method of Claim 29, wherein the mixture is cooled at between 0 degree  and -5 degree C for between 5 and 12 minutes. 31. The method of Claim 24, wherein the diyne-ligand mixture is cooled during polymerization to between 1 degree  and 22 degree C. 32. The method of Claim 31, wherein the diyne-ligand mixture is cooled during polymerization to between 16 degree -19 degree C. 33. The method of Claim 24, wherein the polymerization is achieved by U.V. irradiation using a pen ray lamp or a hand-held lamp. 34. The method of Claim 24, wherein the polymerization is achieved by gamma radiation, electron beam or X-rays, or other low energy ionizing source. 35. The method of Claim 24, wherein the polymerization is accomplished with an energy dose of 10-100 MJ/cm<2>. 36. A method for the direct detection of an analyte in solution, comprising: a) contacting the suspended assay assemblies of Claim 1 with a test sample, and b) observing the solution for a change in color to indicate the presence of an analyte.