Capillary sieving electrophoresis/micellar electrokinetic capillary chromatography for two-dimensional protein fingerprinting of single mammalian cells.
We have developed a two-dimensional capillary electrophoresis method for the study of protein expression in single mammalian cells. The first-dimension capillary contains an SDS-pullulan buffer system to perform capillary sieving electrophoresis, which separates proteins based on molecular weight. The second-dimension capillary contains an SDS buffer for micellar electrokinetic capillary chromatography. After a 6-min-long preliminary separation, fractions from the first capillary are successively transferred to a second capillary, where they undergo further separation by MECC. Over 100 transfers and second-dimension separations are performed over an approximately 3.5-h-long period. We demonstrate this technology by generating protein fingerprints from single native MC3T3-E1 osteoprogenitor cells and MC3T3-E1 cells transfected with the human transcription regulator TWIST. We also present single-cell protein fingerprints from MCF-7 breast cancer cells before and following treatment to induce apoptosis.
Hu
Shen
S
Department of Chemistry, University of Washington, Seattle, Washington 98195, USA.
Michels
David A
DA
Fazal
Md Abul
MA
Ratisoontorn
Chootima
C
Cunningham
Michael L
ML
Dovichi
Norman J
NJ
D000818:Animals; D001943:Breast Neoplasms / Q000473:pathology; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D005260:Female; D006801:Humans; D051379:Mice; D008970:Molecular Weight; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D014157:Transcription Factors / Q000502:physiology; D014407:Tumor Cells, Cultured
D011506:Proteins; D014157:Transcription Factors