Mass mapping of cancer cell lysates using two-dimensional liquid separations, electrospray-time of flight-mass spectrometry, and automated data processing.
Intact protein masses from immortal, nontransformed MCF10A, a human breast epithelial cell line, and its malignant derivative MCF10CA1a.cl1 have been mapped using a combination of all-liquid separations and automated data interpretation. Preparative liquid isoelectric focusing combined with nonporous silica reverse-phase high-performance liquid chromatography allows efficient separation of a large number of proteins in complex mixtures such as whole-cell lysates. Molecular weight determination of these proteins is achieved using electrospray-time of flight-mass spectrometry, however, manual data analysis for these separations is both complex and time-consuming. Protein mass mapping can be significantly enhanced by automating deconvolution functions typically performed manually, with resulting reductions in hands-on analysis time from 20-30 h per chromatogram to approximately 15 min. This reduction in analysis time allows for rapid screening of cancer cell lines for potential biomarkers over a wider pI range than would otherwise be possible.
Buchanan
Nathan S
NS
Department of Chemistry, The University of Michigan, Ann Arbor, MI 48109-1055, USA.
Hamler
Rick L
RL
Leopold
Peter E
PE
Miller
Fred R
FR
Lubman
David M
DM
D014408:Biomarkers, Tumor / Q000032:analysis* / Q000302:isolation & purification; D001943:Breast Neoplasms / Q000737:chemistry*; D002457:Cell Extracts / Q000737:chemistry; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D005260:Female; D006801:Humans; D007525:Isoelectric Focusing; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000032:analysis* / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D014408:Biomarkers, Tumor; D002457:Cell Extracts; D009363:Neoplasm Proteins