Biospecific multianalyte assay method

• US 5,028,545 A
• Filed: 06/09/1988
• Issued: 07/02/1991
• Est. Priority Date: 06/16/1987
• Status: Expired due to Term

• Current Assignee: Wallac Oy (Revvity, Inc.)
• Original Assignee: Wallac Oy (Revvity, Inc.)
• Inventors: Soini, Erkki J.
• Primary Examiner(s): Wax, Robert A.
• Assistant Examiner(s): Sisson, Bradley L.
• Application Number:
  US07/204,258
• Time in Patent Office: 1,118 Days
• Field of Search: 436/501, 435/6, 435/7
• US Class Current: 436/501
• CPC Class Codes: G01N 15/14 Optical investigation techniques, e.g. flow cytometry
• CPC Class Codes: G01N 21/6408 with measurement of decay time, time resolved fluorescence
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form
• CPC Class Codes: G01N 33/582 with fluorescent label

Abstract

In a biospecific multianalyte assay the use of microspheres and fluorescent labels with substantially different fluorescence decay times, is combined. The assay is performed in a suspension of microspheres in the form of a pool of different microsphere categories, where the categories represent different analytes. The microspheres belonging to the respective categories are first coated with a specific reactant, i.e. the microspheres function as a solid support for the reactant and for a biospecific reaction. Fluorescent labels having a short decay time are used to identify the category of each individual microsphere, while fluorescent labels having a long decay time are used to determine the concentration of a particular analyte on the microsphere by means of the biospecific reaction.

Claims

1. Biospecific multianalyte assay method, characterized by preparing categories of microspheres representing different analytes to be assayed, said categories comprising different amounts of a fluorescent substance having a short decay time,

coating each category of microspheres with a biospecific reactant,

pooling the different categories of microspheres together in a suspension,

adding a sample containing analytes to be assayed to the suspension,

adding a mixture of biospecific reactants labelled with a fluorescent compound having a long decay time to the suspension to initiate biospecific reactions between the analytes and the belled reactants and microsphere-associated reactants,

diluting the suspension to reduce the concentration of labelled reactants not bound to the microspheres,

exciting both the fluorescent substance having a short decay time and the fluorescent compound having a long decay time, associated with the microspheres, to generate fluorescence emissions,

converting the fluorescence emissions to electrical signals,

identifying the category of each microsphere on the basis of the strength of the electrical signal resulting from the short decay time fluorescent substance,

measuring the concentration of the analyte on each microsphere on the basis of the strength of the electrical signal resulting from the long decay time fluorescent compound.