Non-specific binding (NSB), especially due to cross-reactivity between specific capture probes and interferents, is a platform-independent issue that limits the detection selectivity of most affinity-based bioanalytical assays. The ability to differentiate and quantify NSB has thus been sought after particularly in label-free biomolecular detections. In this paper, the Langmuir formulation has been applied to understand the transient competitive binding between the target molecules and interferents with the capture probes in complex solutions. By tracking the deviation from the ideal interferent-free case, two NSB differentiation approaches have been proposed. The first one requires multiple measurements during the initial transient and has a short turnaround time. The second approach necessitates only one transient and one equilibrium sampling, which offers a better selectivity over the first approach. The applicability of each approach has been scrutinized depending on whether the interferents would contribute a signal comparable to that of the targets. Finally, the achievable detection selectivity has been correlated to the intrinsic sensitivity of the bioanalytical assay.

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