Objective To explore the methods and mesures for the pre-analytical quality control of test specimen and to improve the accuracy and reliability of test results.Methods According to the relevant requirements of IS015189,various measures for the specimen circulation process and collection technology two aspects were taken to control the full links of clinical specimen collection and transport,and the incidence of unqualified specimen before and after the improvements were analyzed.Results After the imple-mentation of the improvement measures,the incidence of unqualified specimens was decreased significantly.Conclusion Implemen-tation of the whole aspect of pre-analytical quality control can effectively improve the quality of specimen in order to improve the test quality.

OBJECTIVETo establish a cell line with human NADH-cytochrome b5 reductase (b5R) deficiency via RNA interference (RNAi).

METHODSTwo siRNA expressing vectors targeting the b5R mRNA were designed and constructed. Hepatocellular carcinoma BEL-7402 cells were transiently transfected with the two recombinants by lipofectamine (TM) 2000, and semi-quantitative RT-PCR was carried out to analyze the suppression of b5R mRNA; BEL-7402 cells stably transfected with the two siRNA expressing vectors were selected in the media with G418. By analyses of the mRNA, enzymatic activity and protein level of b5R, several cell clones with deficiency of b5R were established. The cell growth curve of BEL-7402 cells with b5R deficiency was detected by MTT assay.

RESULTSTwo siRNA expressing vectors targeting b5R mRNA were obtained, namely pSib5R-1 and pSib5R-2. When BEL-7402 cells were transfected transiently with pSib5R-2, the expression of b5R mRNA was significantly suppressed with a suppression ratio of 68.3%, indicating that pSib5R-2 could trigger the degradation of b5R mRNA effectively. Eighteen clones stably integrated exogenous plasmids were obtained. In two clones from pSib5R-2 transfection, the expression of b5R mRNA was suppressed by up to 48.2% and 56.2%, and the enzymatic activity was inhibited by up to 54.6% and 63.5%, respectively. The protein levels also decreased significantly. The defect of b5R did not change the cell growth rate.

CONCLUSIONThe expression of b5R in BEL-7402 could be suppressed by vector-based RNA interference effectively. We established a cellular model with defect of b5R successfully, which can be used as a tool in investigation of the biological function of b5R and molecular mechanism of type II recessive congenital methemoglobinemia.

Cell Culture Techniques ; Cell Line, Tumor ; enzymology ; Cells ; enzymology ; Cytochrome-B(5) Reductase ; deficiency ; genetics ; Gene Expression ; drug effects ; Genetic Vectors ; Humans ; RNA Interference ; physiology ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; pharmacology ; Transfection