The expanding use of surfactants for proteome sample preparations has prompted 
the need to systematically optimize the application and removal of these 
MS-deleterious agents prior to proteome measurements. Here we compare four 
detergent cleanup methods (trichloroacetic acid (TCA) precipitation, 
chloroform/methanol/water (CMW) extraction, a commercial detergent removal spin 
column method (DRS) and filter-aided sample preparation (FASP)) to provide 
efficiency benchmarks with respect to protein, peptide, and spectral 
identifications in each case. Our results show that for protein-limited samples, 
FASP outperforms the other three cleanup methods, while at high protein amounts, 
all the methods are comparable. This information was used to investigate and 
contrast molecular weight-based fractionated with unfractionated lysates from 
three increasingly complex samples ( Escherichia coli K-12, a five microbial 
isolate mixture, and a natural microbial community groundwater sample), all of 
which were prepared with an SDS-FASP approach. The additional fractionation step 
enhanced the number of protein identifications by 8% to 25% over the 
unfractionated approach across the three samples.