A magnetic bead-based serum proteomic fingerprinting method for parallel analytical analysis and micropreparative purification.
ProteinChip surface-enhanced laser desorption/ionization technology and magnetic beads-based ClinProt system are commonly used for semi-quantitative profiling of plasma proteome in biomarker discovery. Unfortunately, the proteins/peptides detected by MS are non-recoverable. To obtain the protein identity of a MS peak, additional time-consuming and material-consuming purification steps have to be done. In this study, we developed a magnetic beads-based proteomic fingerprinting method that allowed semi-quantitative proteomic profiling and micropreparative purification of the profiled proteins in parallel. The use of different chromatographic magnetic beads allowed us to obtain different proteomic profiles, which were comparable to those obtained by the ProteinChip surface-enhanced laser desorption/ionization technology. Our assays were semi-quantitative. The normalized peak intensity was proportional to concentration measured by immunoassay. Both intra-assay and inter-assay coefficients of variation of the normalized peak intensities were in the range of 4-30%. Our method only required 2 microL of serum or plasma for generating enough proteins for semi-quantitative profiling by MALDI-TOF-MS as well as for gel electrophoresis and subsequent protein identification. The protein peaks and corresponding gel spots could be easily matched by comparing their intensities and masses. Because of its high efficiency and reproducibility, our method has great potentials in clinical research, especially in biomarker discovery.
Wong
Melody Y M
MY
Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong SAR, PR China.
Yu
Karen O Y
KO
Poon
Terence C W
TC
Ang
Irene L
IL
Law
Man-Ki
MK
Chan
Kiwi Y W
KY
Ng
Eddy W Y
EW
Ngai
Sai-Ming
SM
Sung
Joseph J Y
JJ
Chan
Henry L Y
HL
D001798:Blood Proteins / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008280:Magnetics; D008863:Microspheres; D010449:Peptide Mapping / Q000379:methods*; D040081:Protein Array Analysis; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001798:Blood Proteins