Multiplexed Proteomic Reactor for the Processing of Proteomic Samples

We report the development of a 96-well plate proteomic
reactor for gel-free processing of minute amounts of
complex proteomic samples. The device performs multiplexed trapping, enrichment, and biochemical processing
of proteins, resulting in concentrated peptide solutions
ready for mass spectrometric analysis. Individual wells
on the reactor can process up to 2 μg of protein. We also
report the coupling of the plate proteomic reactor with
protein fractionation using size-exclusion chromatography
for large-scale identification of proteins. To illustrate the
potential of this approach, we separated 400 μg of MCF7
cell lysate using size-exclusion chromatography and processed 35 protein fractions on the reactor plate. Using
stringent criteria when searching the data, a total of 875
unique proteins were identified. More relaxed searching
conditions associated with a 1% false positive rate led to
the identification of 2683 unique proteins, meaning that
one protein was identified per 3−10 ng of total protein
lysate loaded on the reactor plate.