同位素示踪微量法测定白血病细胞DNA-甲基转移酶活性  

Detection of DNA-methyltransferase activity of leukemia cells with radiology microassay

作　　者：白志勇[1] 徐国宾[2] 武淑兰[1] 

机构地区：[1]北京医科大学第一医院血液科 [2]北京医科大学第一医院检验科

出　　处：《北京医科大学学报》2000年第1期76-78,共3页Journal of Peking University(Health Sciences)

基　　金：卫生部科学研究基金! ( 94 1 2 5 0 )

摘　　要：目的:建立3H标记同位素示踪微量分析法用于白血病细胞MTase活性测定,并评价其准确性、敏感性及可靠性;了解急性白血病患者骨髓细胞MTase活性改变的规律。方法:以Poly[d(IC)d(IC)]为甲基受体,3HSAM为甲基供体,测定骨髓单个核细胞和白血病细胞系的MTase活性。以细胞裂解蛋白作用2h所得的每分计数值表示酶活性。每次测定设不加Poly[d(IC)d(IC)]的阴性对照,以HL60细胞MTase活性为100%,所测样本值均和其相比,以相对数表示结果。结果:当细胞裂解蛋白质量在0.25～7.5μg之间时,本法线性良好,Y每分计数=2864X+142(r=0.998,P<0.01)。方法灵敏度为2.4×104Bq,批间变异系数11.4%(n=9),批内变异系数5.9%(n=4)。15例急性白血病MTase活性平均值为(9.1±4.6)%,较11例非恶性血液病对照组[平均(2.1±1.6)%]明显升高。11例缓解期急性白血病MTase活性平均(4.1±2.6)%,和对照组无明显区别。结论:同位素微量分析法测定MTase活性灵敏度高、重复性好。MTase活性增高是急性白血病的高发现象。急性白血病缓解后MTase活性下降。MTase活性测定可能对监测白血病病情变化有一定意义。Objective:To establish a sensitive radiology microassay technique for studying activity of DNA methyltransferase in leukemia cells, and evaluate its feasibility. Methods: 1.1×10 8 Bq 3H SAM used as a methyl donor and 0.5 μg Poly[d(I C)d(I C)] as a receptor, MTase activity in mononucleated cells from bone marrow of patients with acute leukemia and of control subjects with no hematological malignancies were detected. cpm were counted with 5 μg cellular protein incubated for 2 hours. All detection was performed in duplicates, each experiment had positive control with HL 60 cells and negative control without Poly [d(I C)d(I C)]. Results: The method was linear up form 700 to 20 000 cpm when the quality of cellular proteins was between 0.25 μg and 7.5 μg, Y cpm = 2 864 X + 142, r =0.998.Within group and day to day variations were 5.9% and 11.4% respectively. The sensitivity of the method was 2.4×10 4 Bq. Mean MTase activity in 15 cases with acute leukemia was (9.1±4.6)%,4 fold higher than that in 11 cases with no hematological malignancies which was ( 2.1± 1.6)% ( P <0.05); and the value from 11 acute leukemia patients in complete remission was ( 4.1± 2.6)%, which was in the same range as the control group. Conclusion:Radiology microassay had high sensitivity and good replication for detecting MTase activity of leukemia cells. MTase activity of patients with acute leukemia increased, and it decreased in thesame range as control group when thecomplete remission was achieved.

关 键 词：白血病 同位素标记 DNA 甲基转移酶 

分 类 号：R733.7[医药卫生—肿瘤]