State Key Laboratory of Supramolecular Structure and Materials, College of Chemistry, Jilin University, Changchun 130012, P. R. China.
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, P. R. China.
Anal Chem. 2022 May 3;94(17):6591-6598. doi: 10.1021/acs.analchem.2c00695. Epub 2022 Apr 21.
Despite recent advances in single-cell analysis techniques, the ability of single-cell analysis platforms to track specific cells that secreted cytokines remains limited. Here, we report a microfluidic droplet-based fluorescence imaging platform that can analyze single cell-secreted vascular endothelial growth factor (VEGF), an important regulator of physiological and pathological angiogenesis, to explore cellular physiological clues at the single-cell level. Two kinds of silica nanoparticle (NP)-based immunoprobes were developed, and they were bioconjugated to the membrane proteins of the probed cell surface the bridging of secreted VEGF. Thus, an immunosandwich assay was built above the probed cell fluorescence imaging analysis of each cell in isolated droplets. This analytical platform was used to compare the single-cell VEGF secretion ability of three cell lines (MCF-7, HeLa, and H8), which experimentally demonstrates the cellular heterogeneity of cells in secreting cytokines. The uniqueness of this method is that the single-cell assay is carried out above the cell of interest, and no additional carriers (beads or reporter cells) for capturing analytes are needed, which dramatically improves the availability of microdroplets. This single-cell analytical platform can be applied for determining other secreted cytokines at the single-cell level by changing other immune pairs, which will be an available tool for exploring single-cell metabonomics.
尽管单细胞分析技术近年来取得了进展，但单细胞分析平台跟踪分泌细胞因子的特定细胞的能力仍然有限。在这里，我们报告了一种基于微流控液滴的荧光成像平台，该平台可分析分泌血管内皮生长因子（VEGF）的单个细胞，VEGF 是生理和病理血管生成的重要调节剂，以探索单细胞水平的细胞生理线索。开发了两种基于二氧化硅纳米颗粒（NP）的免疫探针，并将其与被探测细胞表面的膜蛋白连接，从而构建了被探测细胞上方的免疫夹心测定法，用于对分离液滴中的每个细胞进行荧光成像分析。该分析平台用于比较三种细胞系（MCF-7、HeLa 和 H8）的单个细胞 VEGF 分泌能力，实验证明了细胞因子分泌细胞的细胞异质性。该方法的独特之处在于单细胞测定是在感兴趣的细胞上方进行的，不需要用于捕获分析物的额外载体（珠子或报告细胞），这极大地提高了微滴的可用性。通过改变其他免疫对，这种单细胞分析平台可用于在单细胞水平上测定其他分泌细胞因子，这将成为探索单细胞代谢组学的有效工具。
