Robust Microflow LC-MS/MS for Proteome Analysis: 38 000 Runs and Counting

Microflow liquid chromatography tandem
mass spectrometry (μLC–MS/MS)
is becoming a viable alternative to nanoflow LC–MS/MS for the
analysis of proteomes. We have recently demonstrated the potential
of such a system operating with a 1 mm i.d. × 150 mm column and
at a flow rate of 50 μL/min for high-throughput applications.
On the basis of the analysis of ∼38 000 samples measured
on two instruments over the past two years, we now show that the approach
is extremely robust. Up to 1500 analyses were performed within one
month, and >14 000 samples could be analyzed on a single
column
without loss of chromatographic performance. Samples included proteomes
of cell lines, tissues, and human body fluids, which were analyzed
with or without prior peptide fractionation or stable isotope labeling.
We show that the μLC–MS/MS system is capable of measuring
2600 proteins from undepleted human plasma and ∼5000 proteins
from crude human urine in 1 day, demonstrating its potential for in-depth
as well as high-throughput clinical application.