On-plate digestion using a commercial microfraction collector for nano-HPLC matrix-assisted laser desorption/ionization tandem time-of-flight protein analysis.
A new method for on-plate protein digestion and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry analysis is proposed involving an automated one-step sample separation using nanoflow HPLC followed by nanoliter fraction collection and on-plate digestion with trypsin. This procedure uses a commercial automatic nanoliter fraction collection system for on-line spotting of the eluent onto a MALDI target. After protein digestion, the reaction is stopped by the addition of acidified matrix using the same automated system. Collected spots are subsequently analyzed using a MALDI tandem time-of-flight (TOF/TOF) mass spectrometer for protein sequencing and identification.
Department of Chemistry, University of Aveiro, 3810-193 Aveiro, Portugal. rvitorino@ua.pt
Rui
R
Vitorino
Sofia
S
Guedes
Kenneth
K
Tomer
Pedro
P
Domingues
José
J
Duarte
Francisco
F
Amado
D053000:Analytic Sample Preparation Methods / Q000379:methods*; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D011506:Proteins / Q000032:analysis* / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D014357:Trypsin / Q000378:metabolism
D011506:Proteins; D014357:Trypsin