On-target digestion of collected bacteria for MALDI mass spectrometry.
An on-target protein digestion system was developed for the identification of microorganisms in collected bioaerosols using off-line matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Bacteria analysis techniques based on MALDI-MS were adapted for use with an orthogonal MALDI quadrupole-time-of-flight mass spectrometer. Bioaerosols were generated using a pneumatic nebulizer and infused into a chamber for sampling. An Andersen N6 single-stage impactor was used to collect the bioaerosols on a MALDI target. On-target digestion was carried out inside temporary mini-wells placed over the impacted samples. The wells served as miniature reactors for proteolysis. Collected test aerosol particles containing the protein cytochrome c and E. coli bacteria were proteolyzed in situ using trypsin or cyanogen bromide. A total of 19 unique proteins were identified for E. coli. Using the TOF-MS spectra of the digested samples, peptide mass mapping was performed using the MASCOT search engine and an iterative search technique.
Dugas
Alton J
AJ
Department of Chemistry, Louisiana State University, Baton Rouge, LA 70803, United States.
Murray
Kermit K
KK
D000336:Aerosols; D001419:Bacteria / Q000302:isolation & purification* / Q000378:metabolism*; D003488:Cyanogen Bromide / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D004926:Escherichia coli / Q000302:isolation & purification / Q000378:metabolism; D010449:Peptide Mapping; D040901:Proteomics; D012997:Solvents / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000378:metabolism
D000336:Aerosols; D012997:Solvents; D045304:Cytochromes c; D014357:Trypsin; D003488:Cyanogen Bromide