Gold nanoparticles enhance fluorescence signals by flow cytometry at low antibody concentrations

Abstract

Flow cytometry is a universally applied technique in many biological and clinical assays to evaluate cells, bacteria, parasites, and particles at a micrometre scale. More advanced flow cytometers can detect small molecules down to the nanometre scale that may identify intracellular nanostructures. Advancements in the field of nanobiotechnology have led to techniques that allow the study of cellular behaviour after exposure to nanomaterials, particularly, metal nanoparticles. The optical properties of gold nanoparticles regarding surface plasmon resonance (SPR) are established to increase the fluorescence quantum yields of several dyes working as optical antennas, enabling the enhancement of light emission in fluorescent emitters. In this work we constructed a nanoprobe using gold nanoparticles coated with primary antibody Cetuximab. Then, we investigated whether this nanoprobe labelled with secondary fluorescent antibody Alexa Fluor 488, at low concentrations, could promote fluorescent signal enhancement, associated with SPR, and detected by the flow cytometry technique. Our results showed an enhanced fluorescent signal likely due to the proximity between the extinction coefficient of gold nanoparticles and the emission peak of Alexa Fluor 488, at exceptionally low concentrations, occurring within a high level of specificity. Moreover, the nanoprobe did not alter the cellular viability suggesting gold nanoparticles as a feasible approach for cell labelling using low concentrations of secondary antibodies for routine flow cytometry applications.