posted on 2022-02-01, 17:11authored byChao Dong, Luke T. Richardson, Touradj Solouki, Kermit K. Murray
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 mm2 area of ablated tissue. Areas of tissue ranging
from 0.01 to 1 mm2 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.