Infrared Laser Ablation Microsampling for Small Volume Proteomics

Abstract

Infrared (IR) laser ablation was used to remove localized tissue regions from which proteins were extracted and processed with a low volume sample preparation workflow for bottom-up proteomics by liquid chromatography tandem mass spectrometry (LC-MS/MS). A polytetrafluoroethylene (PTFE) coated glass slide with 2 mm diameter microwells was used to capture ablated rat brain tissue for in situ protein digestion with submicroliter solution volumes. The resulting peptides were analyzed with LC-MS/MS for protein identification and labelfree quantification. The method was used to identify an average of 600, 1350, and 1900 proteins from ablation areas of 0.01, 0.04, and 0.1 mm 2 , respectively, from a 50 μm thick rat brain tissue section. Differential proteomics of 0.01 mm 2 regions captured from cerebral cortex and corpus callosum was accomplished to demonstrate the capabilities of the approach.

Direct 3D Sampling of the Embryonic Mouse Head: Layer-wise Nanosecond Infrared Laser (NIRL) Ablation from Scalp to Cortex for Spatially Resolved Proteomics

“…A novel approach to overcome the above-mentioned limitations is the nanosecond infrared laser (NIRL) for a fast simultaneous tissue sampling and homogenization. − The process is based on tissue irradiation with ultrashort infrared laser pulses at a wavelength of 2940 nm. During this process, the energy is absorbed and converted into vibrational motion of the OH stretch band of water molecules in the cell.…”