Quantitative analyte assay device and method

An assay device for the quantitative determination of an analyte, comprising a test strip containing a porous test membrane allowing for capillary flow of the analyte and complexes of the analyte, a porous upstream membrane in fluid connection with the test membrane and a porous downstream membrane in fluid connection with the test membrane,

• (a) said test membrane comprising (i) a test band site having immobilized thereon a ligand capable of competing with the analyte for binding a label conjugate capable of reacting with the analyte to the test band site, and, (ii) a first standard band site having immobilized thereon a first calibrator agent corresponding to a first known high concentration of the analyte, and a second standard band site having immobilized thereon a second calibrator agent corresponding to a second known low concentration of the analyte, wherein the two calibrator agents are different and are capable of reacting with different label conjugates and binding such to the standard band sites to provide a known label response in the standard band sites,
• (b) said upstream membrane having a site for the application of a sample to be analyzed, and having a site downstream from the sample application site for depositing the label conjugate capable of reacting with the analyte and depositing the two label conjugates capable of reacting with the immobilized calibrator agents in the standard bands, and
• (c) said downstream membrane capable of absorbing said sample and providing the capillary flow for the sample through the upstream and test membrane.