Objective:To clarify further the mechanisms for total saponins of Panax ginseng(TSPG)in inducing differentiation of K562 cells,and to provide the theoretical basis and the experimental evidence of its clinical application.Methods:By using cell culture in vitro,morphological observation and flow cytometry,the effect of TSPG on K562 cells in expressing CD 15 ,HIR 2,EPO-R and GM-CSF-R were studied.Results:The results indicated that TSPG could induce the differentiation of K562 cells toward proerythroid cells and progranulocytic cells.It was also shown that after induction by TSPG,the ratio of positive K562 cells expressing CD 15 ,HIR 2,erythropoietin recepter(EPO-R) and granulocyte-macrophage colony stimulating facter recepter(GM-CSF-R) increased.Conclusion:K562 cells are abnormal cells whose differentiation are blocked,but its biological features are similar to the normal hematopoietic stem cells.The mechanism for TSPG inducing differentiation of K562 cells may be related to the higher expression of EPO-R and GM-CSF-R.

Objective To investigate the effects of total saponins of Panax ginseng(TSPG) combined with erythropoietin(EPO) or granulocyte macrophage colony stimulating factor(GM CSF) on the differentiation of K562 cells. Methods Cells were cultured in vitro . Morphologic changes of cells were observed with light microscope and electron microscope. The levels of hemoglobin, peroxidase, non specific lipase ? NAE in cells were detected respectively by benzidine staining, peroxidase(POX) staining and ? NAE cytochemical staining. Levels of cell surface differentiation antigens CD15, HIR2, EPO R and GM CSF R were determined by immunocytochemistry. Results TSPG combined with EPO or GM CSF resulted in more mature morphological features of K562, increased expressions of hemoglobin and myeolperoxidase in K562 cells, and enhanced expressions of cell surface differentiation antigens CD15, HIR2, EPO R and GM CSF R. Conclusion TSPG combined with cytokine EPO or GM CSF can induce more mature differentiation of K562 cells.

Objective:To prepare monoclonal antibodies(McAb)against cystatin C(Cys C)and to establish the particle enhanced turbidimetric immunoassay(PETIA)for determining human serum Cys C.Methods:The prokaryotic expression vector pET32a(+)/Cys C was constructed and Cys C expression was induced.McAbs against Cys C were prepared with the hybridoma technique after mice were immunized with the purified recombinant protein.Then the McAbs were covalently attached to uniform microparticles,PETIA method for determination of human serum Cys C was established,and primary evaluation tests of methodology were performed.Results:Three hybridoma cell lines were obtained successfully,the secreted antibodies were isotype of IgG1,and Western blot confirmed that the antibodies reacted specifically to the Cys C protein.After one of the hybridoma cell lines was injected into mice abdominal cavity,the ascites abundant for McAb was obtained.The titer of the McAb against the purified protein was 1∶4?106.With the self-made McAb,PETIA for human serum Cys C was established.The primary evaluation tests of methodology revealed that self-established PETIA method had a satisfactory performance,which was equal to the import kit.Conclusion:The prepared McAb against Cys C is prepared,which could be used to establish PETIA for determining human serum Cys C.

Objective To investigate the clinical application of Cystatin C as a biological marker for monitoring hepatic pathological change in patients with virus hepatitis.Methods Two hundred and seven patients infected with hepatitis B or C virus(HBV, HCV)were divided into cirrhosis group(group A),chronic HBV group(group B),chronic HCV group(group C),and liver cancer group(group D). 32 healthy controls(group H) were recruited . The serum TIMP-1,TIMP-2,and Cystatin C as well as some traditional markers for monitoring liver function and renal function including creatinine, creatinine clearance rate, alanine transaminase, and aspartate transaminase were determined.Results In these groups, serum Cystatin C(F=28.334, P

Object To clarify the mechanism for total saponins of Panax ginseng C.A.Mey. (TSPG) inducing K562 cells to apoptosis, and to provide the theoretical basis and the experimental evidence of TSPG's clinical application. Methods By using in vitro cell culture, morphometry, flow cytometry, morphological investigation and immunocytochemistry, the effects of TSPG on apoptosis in K562 cells were studied. Results The results indicated that TSPG could inhibit the proliferation of K562 cells significantly, and induce K562 cells to apoptosis. It was also showed in the experiments that after induced by TSPG, the ratio of positive K562 cells expressing C-MYC and BCL-2 is decreased. Conclusion The mechanism for TSPG to induce K562 cells to apoptosis may be related to the expression of oncogene in K562 cells.

Objective To construct a prokaryotic expression vector of cystatin C (Cys C), purify Cys C protein produced by the expression system, and prepare its antiserum. Methods Total RNA was isolated from HL-60 cells, and human Cys C gene was amplified with RT-PCR. The cDNA fragment was cloned into pMD18-T vector and which was confirmed by sequencing. The enzyme-digested target fragment was cloned into PET-32(a) expression vector and transfected into E.coli. BL 21(DE3), in which Cys C expression was induced. After the inclusion body protein was purified through Ni2+ affinity chromatography, processed by dialysis, identified by Western blotting, a rabbit was immunized with the fusion protein, and the antiserum was obtained. Results The result of DNA sequence analysis showed that the cloned Cys C gene sequence was completely corresponding to GenBank data. SDS-PAGE and Western blotting showed that the expressed Cys C fusion protein was about 35?103, mainly existing in the inclusion body of E.coli., that could be purified through Ni2+ affinity chromatography. The titer of the antiserum to the purified protein was 1∶8 000 by ELISA, and Western blotting confirmed that the antiserum reacted specifically to the Cys C protein. Conclusion A recombinant Cys C protein and the specific polyclonal antibody have been obtained, which provides a basis for establishment of immunoassays of human Cys C.

Aiming at the malpractice of traditional education mode,we have carried out a series of experimental teaching reform on laboratory immunology,including developing compositive and devised experiments,applying the fruit of scientific research into experimental teaching,doing the special discuss after immune experiments as well as going to clinical laboratory to learn termly,which can adequately embody the students' principle part and the teachers' leading role,and improve the students' comprehensive qualities.

Objective To establish a continuous monitoring assay of serum argininosuccinate lyase (ASL) activity with automatic biochemistry analyzer, and perform methodology validation and preliminary clinical application.Methods According to the chemical reaction catalyzed by ASL and the working characteristics of automatic biochemistry analyzer, an enzyme coupled reaction system with high specificity was set up, and the methodology validation was performed.Three hundred and nine patients with various liver diseases, 269 non-liver disease patients and 40 healthy controls were enrolled in this study.Serum ASL, ALT, and AST level were determined in all subjects.Results A new kinetics assay of ASL activity was set up with automatic biochemistry analyzer.The methodological validation demonstratod that inter-assay and intra-assay coefficient of variation were 4.0% and 5.9% respectively and the mean recovery was 100.5%.The linear range was 0-167.7 U/L.The lowest detection limit was approximately 0 U/L.The interference test showed that there is no significant interferences while the concentration of bilirubin is less than 342 μmoL/L or commonly used anticoagulants is employed at their routine concentrations.However,interference was significant when Hb level is more than 0.06 g/L.Preliminary study of clinical application showed that there was no significant difference of serum ASL level between non-liver disease group and healthy group ( q = 0.027, P = 0.979 ), but there was significant differences for both serum ALT and AST levels (ALT:q =6.461,P =0.000;AST:q =6.481,P =0.000).Conclusions A continuous monitoring assay for the determination of serum ASL activity is successfully established. Serum ASL may be a good biomarker for liver injury.

In order to meet the education reform and the needs of diversified society,according to the talents training goal of generic eagle plan of Inspection Department of Chongqing Medical University,this article use the pin type education management to awake the student's potential advantage,cultivate a variety of laboratory medicine talented person.This article was focused on pin type education management scheme,implementation methods,results and experience.

AIM: The objectives of the present study were to examine the effect of Jumi(JM) extraction on relaxation of isolated rat aortic rings,and to elucidate its mechanisms.METHODS: The thoracic aortic rings with and without endothelium of male Sprague-Dawley rats were mounted on a bath system.Vasodilatation of aortic rings preconstricted with 10-6 mol/L of phenylephrine(PE) was measured.RESULTS: JM extraction(0.5-8 g/L) caused a concentration-dependent relaxation in aortic rings.The extent of relaxation was larger in endothelium-intact aortic rings than that in endothelium-denuded aortic rings.Both L-NAME [a nitric oxide synthase(NOS) inhibitor] and high potassium(20 mmol/L KCl) partly abolished the relaxation action of JM extraction in endothelium-intact aortic rings.Pretreatment with L-NAME also inhibited the relaxation response to JM extraction in endothelium-denuded aortic rings.After incubation with JM extraction,NOS activities enhanced both in endothelium-intact and endothelium-denuded aortic rings.CONCLUSION: JM extraction causes relaxation of aortic rings through endothelium-dependent and independent pathways.The mechanisms might be involved in NOS and endothelium-derived hyperpolarizing factor.