Mass spectrometric compatibility of Deep Purple and SYPRO Ruby total protein stains for high-throughput proteomics using large-format two-dimensional gel electrophoresis.
In order to identify putative biomarkers from two-dimensional (2D) gel electrophoresis it is necessary to use a visualization technique that is sensitive, has a large dynamic range and does not interfere with the identification of the protein. As mass spectrometry increases in sensitivity more pressure is placed on visualization techniques that facilitate proteomic workflows but do not interfere with downstream processing. Two stains reported to meet these requirements are SYPRO Ruby (Invitrogen) and Deep Purple (GE Healthcare). This study examined the compatibility of these stains with protein identification by selecting spots from replicate 2D gels of human plasma and subjecting these to protein identification using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Using a test of two populations of proportions it was found that proteins were statistically more likely to be identified from gels stained with Deep Purple. Additionally, the identifications from Deep Purple stained gels are of higher quality because they are based on multiple peptides.
Nock
Christina M
CM
GlaxoSmithKline Pharmaceuticals, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, UK.
Ball
Malcolm S
MS
White
Ian R
IR
Skehel
J Mark
JM
Bill
Louisa
L
Karuso
Peter
P
D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D009942:Organometallic Compounds / Q000737:chemistry*; D010449:Peptide Mapping / Q000379:methods*; D040081:Protein Array Analysis / Q000379:methods*; D040901:Proteomics / Q000379:methods*
D009942:Organometallic Compounds; C515601:Sypro Ruby