CyDye immunoblotting for proteomics: co-detection of specific immunoreactive and total protein profiles.
The development of ECL-Plex CyDye-conjugated secondary antibodies allows the advancement of conventional Western blotting, opening up possibilities for highly sensitive and quantitative protein confirmation and identification. We report a novel proteomic method to simultaneously visualise the total protein profile as well as the specific immunodetection of an individual protein species by combining cyanine CyDye pre-labelled proteins and antibody immunoblotting. This technique proposes to revolutionise both 2-D immunoprobing and protein confirmation following MS analysis.
Donoghue
Pamela M
PM
Proteome Research Centre, UCD Conway Institute of Biomolecular and Biomedical Research, Dublin, Ireland.
McManus
Ciara A
CA
O'Donoghue
Niaobh M
NM
Pennington
Stephen R
SR
Dunn
Michael J
MJ
D000595:Amino Acid Sequence; D002232:Carbocyanines; D005456:Fluorescent Dyes; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D007120:Immunochemistry; D018832:Molecular Chaperones; D009206:Myocardium / Q000737:chemistry; D009363:Neoplasm Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D040081:Protein Array Analysis / Q000379:methods; D011506:Proteins / Q000276:immunology / Q000302:isolation & purification; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D002232:Carbocyanines; D005456:Fluorescent Dyes; D055551:HSP27 Heat-Shock Proteins; C057725:HSPB1 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D011506:Proteins; C089187:cyanine dye 3; C085321:cyanine dye 5