Purification of tryptic peptides for mass spectrometry using polyvinylidene fluoride membrane.
A simple procedure for the purification of tryptic peptides, prior to mass spectrometric analysis, using polyvinylidene fluoride membrane (PVDF) is described. The sensitivity of mass spectrometric analysis is such that minor impurities in tryptic peptide digests suppress the signal obtained. However, we obtained useful signal, from a sample that did not yield any spectra earlier, by purifying the sample using PVDF membrane. For this, the tryptic peptide digest was first spotted on the membrane which was then air-dried and washed. Further, the membrane was extracted with trifluoroacetic acid (TFA) and acetonitrile and subjected to mass spectrometric analysis. This procedure enabled us to identify a cross-reactive D1 antigen on the neutrophil surface that bound antibodies that targeted 60 kD Ro autoantigen in systemic lupus erythematosus, an autoimmune disorder.
Kurien
B T
BT
Arthritis and Immunology Program, Oklahoma Medical Research Foundation, Oklahoma City 73104, USA. kurienb@omrf.ouhsc.edu
Matsumoto
H
H
Scofield
R H
RH
D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008567:Membranes, Artificial; D009504:Neutrophils / Q000737:chemistry*; D010455:Peptides / Q000032:analysis / Q000302:isolation & purification*; D011145:Polyvinyls / Q000737:chemistry*; D014357:Trypsin / Q000378:metabolism*; D005465:Fluorocarbon Polymers
D008567:Membranes, Artificial; D010455:Peptides; D011145:Polyvinyls; D014357:Trypsin; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers