BIOLOGICAL LIGAND-RECEPTOR BINDING ASSAYS USING SPATIALLY RESOLVED DUAL ILLUMINATION DIGITAL IMAGE ANALYSIS TO ELIMINATE A WASHING STEP

摘要

A method of detecting a ligand in a sample comprising the steps of: (a) performing a ligand-receptor binding assay, wherein the ligand-receptor binding assay comprises combining a receptor attached to a solid support, a conjugate comprising a label, and a sample suspected of containing a ligand in a reaction vessel and allowing a reaction to occur; (b) acquiring an image to determine the location of the conjugate; (c) selecting at least one region of interest from the image; and (d) detecting a ligand in the sample. In particular, the detecting step comprises (i) selecting pixels from the at least one region of interest for analysis; (ii) calculating and recording the average and variance of the counts per pixel for the pixels selected in step (i); (iii) omitting pixels that have counts greater than or less than a specified variance; and (iv) calculating average counts per pixel of the remaining pixels.