Purification of Tryptic Digests on Polyvinylidene Difluoride Membrane

Minor impurities in tryptic peptide digests can affect the signal obtained in matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Therefore, it becomes necessary to purify the digests, especially those that fail to yield good mass spectra. Here, we describe a simple protocol using polyvinylidene difluoride membrane for purifying tryptic peptides prior to mass spectrometric analysis. The tryptic digest is spotted on a polyvinylidene difluoride membrane, air-dried, and washed. The membrane is then extracted with trifluoroacetic acid/acetonitrile and the extract is then subjected to matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This method enabled us to identify a cross-reactive D1 autoantigen on the surface of neutrophils that bound antibodies targeting Ro 60 autoantigen in systemic lupus erythematosus.