title: [Detecting phospho-signaling protein of bone marrow leukemia cells by phospho-signaling flow cytometry].
abstract: The purpose of this study was to establish the phospho-specific flow cytometry (phospho-flow) to detect the phosphorylated signaling proteins of leukemia cells and to evaluate its useful value in leukemia study. The bone marrow of leukemia children was collected, and treated by phospho-flow of extracted mononuclear cells (MNC) and phospho-flow of directly fixed bone marrow (BM) respectively. In phospho-flow of extracted MNC, the MNC extracted from BM were fixed and permeabilized, then were cultured with P-AKT and P-ERK1/2, finally were analyzed by flow cytometry. In phospho-flow of directly fixed BM, the BM was treated with fixation/lysis buffer and 90% methanol, then were incubated with the surface CD antibody, P-AKT and P-ERK1/2, finally the treated BM cells were analyzed by flow cytometry. The results showed that the positive rates of P-AKT and P-ERK1/2 in MNC treated by phospho-flow of extracted MNC of 26 leukemia children were 46.2% and 30.8% respectively, while the positive rates of P-AKT and P-ERK1/2 in BM treated by phospho-flow of directly fixed BM were 50.0% and 38.5% respectively. The comparison of positive rates of P-AKT and P-ERK1/2 between the 2 treatment protocol showed no difference (p > 0.05). It is concluded that the phospho-flow of directly fixed BM established by our laboratory can be used to analyze the signaling proteins of leukemia cells.
authors: Department of Pediatrics, Guangzhou Medical College, Guangzhou, Guangdong Province, China.
authors: Hua-Mei
authors: HM
authors: Li
authors: Yi-Ru
authors: YR
authors: Zeng
authors: Fu-Xiong
authors: FX
authors: Chen
authors: Li-Ping
authors: LP
authors: Wu
authors: Feng-Gui
authors: FG
authors: Wei
authors: Jia
authors: J
authors: Tao
authors: Hui-Min
authors: HM
authors: Lu
authors: Zi-Liang
authors: ZL
authors: Wu
mesh_terms: D001853:Bone Marrow / Q000378:metabolism*; D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism*; D002648:Child; D002675:Child, Preschool; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D007223:Infant; D007938:Leukemia / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D008297:Male; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D051057:Proto-Oncogene Proteins c-akt