Discovery proteomics by mass spectrometry comparing secreted proteins from young and old mouse lung mesenchymal stromal cells

Lung mesenchymal stromal cells (L-MSCs) were isolated from the young ( 3
months) and old (22-24 months) mice (N = 4 each) following collagenase
digestion of the lungs and anchorage-dependent growth. Culture media was
collected from young and old L-MSCs (10^6 cells) grown in culture
for 24 hrs in serum-free condition. Culture media was then 
centrifuged to remove any cellular debris. Five (5) ml of supernatants
were concentrated using 3kDa Molecular weight cut-off filters, and 10
micrograms of proteins were run as multiple MW 1D PAGE fractions
using a standard GeLC approach with a nano-HPLC in-line with a Velos Pro
Orbitrap MS. Individual data files were searched using Sequest,
the results were then combined for each sample, followed by grouping,
filtering, and quantifying by normalized spectral counts.