Single-cell lysis for visual analysis by electron microscopy

Abstract

The stochastic nature of biological systems makes the study of individual cells a necessity in systems biology. Yet, handling and disruption of single cells and the analysis of the relatively low concentrations of their protein components still challenges available techniques. Transmission electron microscopy (TEM) allows for the analysis of proteins at the single-molecule level. Here, we present a system for single-cell lysis under light microscopy observation, followed by rapid uptake of the cell lysate. Eukaryotic cells were grown on conductively coated glass slides and observed by light microscopy. A custom-designed microcapillary electrode was used to target and lyse individual cells with electrical pulses. Nanoliter volumes were subsequently aspirated into the microcapillary and dispensed onto an electron microscopy grid for TEM inspection. We show, that the cell lysis and preparation method conserves protein structures well and is suitable for visual analysis by TEM.

Keywords: AM; BHK; ECL; EM; ET; Electron microscopy; FEA; FS; HRP; ID; ITO; MS; Microfluidics; OD; OM; PDMS; PEEK; RPPA; SNR; Single-cell analysis; Single-cell lysis; Systems biology; TEM; UA; ammonium molybdate; baby hamster kidney; ddH(2)O; double-distilled water; electron microscopy; electron tomography; enhanced chemiluminescence; finite element analysis; fused silica; horseradish peroxidase; indium tin oxide; inner diameter; mass spectrometry; optical microscope; outer diameter; poly(dimethylsiloxane); poly(ether–ether–ketone); reverse-phase protein arrays; signal-to-noise ratio; transmission electron microscopy; uranyl acetate.