Proposta de mitigação das interações inespecíficas em um biossensor óptico Label Free: aplicação na plataforma BioSearch

Abstract

Non-specific interactions represent one of the main challenges for the application of label-free biosensors: devices capable of detecting biomolecular interactions without the use of secondary markers. In complex biological samples, such as human serum, these interactions, caused by non-selective binding of biomolecules (interferents) to the sensor surface, compromise selectivity, increase background noise, and hinder signal interpretation. In this work, a novel approach is presented for mitigating such interactions using particles referred to as Nanocleansers, which are gold nanorods (AuNbs) with elongated morphology and localized surface plasmon resonance (LSPR) above 800 nm, outside the detection range of the analytical system. The methodology was implemented on the optical BioSearch platform and validated using biosensors functionalized with the N protein of SARS-CoV-2. The detection mechanism of the platform is based on changes in the dynamic depolarized light scattering (DDLS) of biosensors in solution, caused by the binding of biomolecules to them. To quantify such changes, the Brownian motion of the biosensors is tracked through fluctuations in scattered light intensity - enhanced by the optical phenomenon of LSPR from AuNbs. In the proposed system, the variation in depolarized scattered light intensity is mainly due to changes in the rotational diffusivity of the particles triggered by binding events at the biosensor surface. These interactions can occur non-specifically with interferents present in real samples, directly compromising the accuracy and sensitivity of the analytical method. To address this challenge, a protocol for mitigating non-specific interactions is proposed, consisting of the prior addition of Nanocleansers to the sample, enabling the sequestration of interferents before they interact with the biosensors - essentially cleaning the sample. Experiments were conducted using both negative and positive serum samples to evaluate the efficacy of the Nanocleansers under different concentrations and physicochemical conditions. Experimental results demonstrated that this strategy significantly reduces the occurrence of non-specific interactions without impairing the specific detection of the target analyte. The proposed approach is effective, simple, and compatible with real matrix applications, representing an advancement in the sensitivity and accuracy of the label-free optical platform: BioSearch.