Departamento de Bioquímica e Imunologia, Universidade Federal de Minas Gerais, Brazil.
Núcleo de Doenças Infecciosas, Universidade Federal do Espírito Santo, Brazil.
J Mater Chem B. 2021 Feb 7;9(5):1414-1423. doi: 10.1039/d0tb02309d. Epub 2021 Jan 19.
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.
流式细胞术是一种广泛应用于许多生物学和临床分析的技术，用于评估细胞、细菌、寄生虫和微尺度的颗粒。更先进的流式细胞仪可以检测到纳米尺度的小分子，这些小分子可以识别细胞内的纳米结构。纳米生物技术领域的进步带来了一些技术，这些技术允许在暴露于纳米材料后研究细胞行为，特别是金属纳米颗粒。金纳米粒子的光学性质，关于表面等离子体共振（SPR），已经确立，以增加几种作为光学天线的染料的荧光量子产率，从而增强荧光发射器的光发射。在这项工作中，我们使用包被有单克隆抗体 Cetuximab 的金纳米粒子构建了一种纳米探针。然后，我们研究了低浓度的这种用二级荧光抗体 Alexa Fluor 488 标记的纳米探针是否能够促进与 SPR 相关的荧光信号增强，并通过流式细胞术技术检测到。我们的结果表明，由于金纳米粒子的消光系数与 Alexa Fluor 488 的发射峰非常接近，在极低的浓度下，出现了高特异性，因此荧光信号增强。此外，该纳米探针没有改变细胞活力，这表明金纳米粒子作为一种可行的方法，用于使用低浓度的二级抗体进行常规流式细胞术应用的细胞标记。
