Cross-talk correction in multiplexing analysis of biological sample

Abstract

Method for determining fluorescence values { [phi]i spe}i[epsilon]{1,2,...,I} of a set of I fluorescent microparticles {[mu]Pi} i[epsilon]{1,2,...,I} of a multiplexed analysis, said microparticles being in a monolayer arrangement, the method comprising: acquiring a digital fluorescence image of the set of fluorescent microparticles {[mu] Pi} i[epsilon]{1,2,...,I} and computing, for each fluorescent microparticle [mu]Pi in the set of fluorescent microparticles {[mu]Pi} i[epsilon]{1,2,...,I}, a fluorescence value [phi]i meas based only on pixels of the acquired image corresponding to said fluorescent microparticle [mu] Pi. The method comprises computing the fluorescence value [phi]i spe of said fluorescent microparticle [mu] Pi by correcting its first fluorescence [phi]i meas by a cross-talkfluorescence contribution [phi]i cross in said first fluorescence [phi]i meas from other fluorescent microparticles {[mu] Pj}ji in the set of fluorescent microparticles {[mu] Pj} i[epsilon]{1,2,...,I}.

Description

technical field [0001] The present invention relates to biological multiplexed assays, in particular to the diagnosis of complex diseases based on the detection of multiple biomarkers in a biological sample. Background technique [0002] The diagnosis and treatment response of complex diseases are often associated with multiple biomolecules rather than a single identifiable biomarker. In contrast to traditional techniques that measure one analyte at a time, multiplexing techniques can measure dozens to thousands of different biomolecules, such as proteins or nucleic acids, from a single biological sample using the same conditions in a single assay. Basically, different types of capture molecules, such as antibodies, target proteins, peptides, or nucleic acids, are placed in an assay device filled with a sample, each capture molecule being designed to form a specific Fluorescently labeled complexes (referred to as "targeted fluorescently labeled biomarkers"). A major challe...

AI Technical Summary

Problems solved by technology

However, existing viable bead-based systems are designed to run cost-effectively in batches and require waiting for enough samples to fill plates, making turnaround time for routine clinical testing slow

Additionally, both techniques suffer from slower binding kinetics as they are primarily driven by diffusion

Even if agitation is used to speed up the process, often this increases the sample incubation time

In addition, diffusion-limited binding mechanisms can lead to intraassay and interassay variability

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