Integrated system for high-throughput protein identification using a microfabricated device coupled to capillary electrophoresis/nanoelectrospray mass spectrometry.
An integrated microsystem providing rapid analyses of trace-level tryptic digests for proteomics application is presented. This modular microsystem includes an autosampler and a microfabricated device comprising a sample introduction port and an array of separation channels together with a low dead-volume facilitating the interface to nanoelectrospray mass spectrometry. Sequential injection and separation of peptide standards and tryptic digests was achieved with a throughput of up to 30 samples per hour with less than 3% sample carryover. Replicate injections of peptide mixtures indicated that reproducibility of migration time was typically better than 2.3% relative standard deviation (RSD) whereas RSD values of 3.7-11.8% were observed on peak height. Mass spectral detection of submicromolar protein digests (< 7 femtomoles/injection) was achieved using a quadrupole/time of flight instrument in less than 2 min/per sample with peak widths of 1.8-7.0 s. The analytical potential of this integrated device for the identification of gel isolated proteins from Neisseria meningitidis immunotype L3 has been demonstrated using both peptide mass-fingerprint database searching and on-line tandem mass spectrometry.
Li
J
J
Institute for Biological Sciences, Ottawa, Ontario, Canada.
Tremblay
T L
TL
Wang
C
C
Attiya
S
S
Harrison
D J
DJ
Thibault
P
P
D000595:Amino Acid Sequence; D019075:Electrophoresis, Capillary / Q000379:methods*; D036103:Nanotechnology; D011506:Proteins / Q000737:chemistry*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D018511:Systems Integration
D011506:Proteins