Infrared Laser Ablation Microsampling with a Reflective Objective.

A Schwarzschild reflective objective with a numerical aperture of 0.3 and working distance of 10 cm was used for laser ablation sampling of tissue for off-line mass spectrometry. The objective focused the laser to a diameter of 5 μm and produced 10 μm ablation spots on thin ink films and tissue sections. Rat brain tissue sections 50 μm thick were ablated in transmission geometry, and the ablated material was captured in a microcentrifuge tube containing solvent. Proteins from ablated tissue sections were quantified with a Bradford assay, which indicated that approximately 300 ng of protein was captured from a 1 mm 2 area of ablated tissue. Areas of tissue ranging from 0.01 to 1 mm 2 were ablated and captured for bottom-up proteomics. Proteins were extracted from the captured tissue and digested for liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis for peptide and protein identification.