Establishment of two-dimensional gel electrophoresis profiles of the             human acute promyelocytic leukemia cell line NB4.
To explore optimum conditions for establishing a two‑dimensional gel electrophoresis             (2-DE) map of the human acute promyelocytic leukemia (APL) cell line NB4 and to             analyze its protein profiles, we extracted total proteins from NB4 cells using             cell disruption, liquid nitrogen freeze-thawing and fracturing by ultrasound,             and quantified the extracted protein samples using Bradford's method. 2-DE was             applied to separate the proteins, which were silver-stained in the gel. Well‑separated             protein spots were selected from the gel using the ImageMaster™ 2D Platinum analysis             system. Moreover, the effects of various protein sample sizes (140, 160 and 180 µg)             on the 2-DE maps of the NB4 cells were determined and compared. Matrix-assisted             laser desorption/ionization time of flight-mass spectrometry (MALDI-TOF-MS), peptide             mass fingerprinting (PMF) and database searching were used to identify the proteins.             When the quantity of loading proteins was 160 µg, clear, well-resolved, reproducible             2-DE proteomic profiles of the NB4 cells were obtained. The average number of             protein spots in 3 gels was 1160±51 with an average matching rate of 81%. A total             of 10 proteins were identified by mass spectrometry and database queries, certain             proteins were products of oncogenes and others were involved in cell cycle regulation             and signal transduction. In summary, 2-DE profiles of the proteome of NB4 cells             were established and certain proteins were identified by MALDI-TOF-MS and PMF             which lay the foundation of further proteomic research of NB4 cells. These data             should be useful for establishing a human APL proteome database.
Department of Hematology, The First Affiliated Hospital, School of Medicine, Xi'an Jiaotong University, Xi'an 710061, PR China.
Pengcheng
P
He
Yanfeng
Y
Liu
Mei
M
Zhang
Xiaoning
X
Wang
Huaiyu
H
Wang
Jieying
J
Xi
Kaihua
K
Wei
Hongli
H
Wang
Jing
J
Zhao
D045744:Cell Line, Tumor; D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism / Q000473:pathology; D010449:Peptide Mapping; D020543:Proteome / Q000032:analysis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D020543:Proteome