Objective To establish a real-time fluorescent quantitative PCR ( FQ-PCR ) method for detection of murine norovirus ( MNV) in laboratory mouse and provide the basis for establishment of a standard detection method for MNV.Methods Specific primers were designed and MNV DNA standards were prepared according to the MNV genome sequences published on NCBI .The specificity, sensitivity, repeatability and stability of the established Q-PCR method were tested.The established Q-PCR method was applied to detect 766 mouse caecum content samples to explore preliminarily the infection status of laboratory mice in Beijing .Results No cross reaction showed in human norovirus and feline calicivirus with the established Q-PCR method.The sensitivity was up to 10 copies/μL.The coefficient of variation ( CV) of intra-assay and inter-assay was less than 2%.There were 301 positive cases detected in the 766 samples of laboratory mice.Conclusions The established FQ-PCR method is accurate and effective with high specificity , sensitivity and repeatabiliy in the quantitative detetion of nucleic acid , and can be applied to rapidly and quantitatively screen MNV in laboratory mice .

Nuclear receptors,the superfamily of ligand-activated transcription factors,play crucial role in the regulation of gene expression of the lipid metabolism,and are involved in many metabolic diseases,such as cholesterol gallstone disease.Recently many domestic and foreign researches on the role of nuclear receptors in the lipid metabolism,help to further elucidate the biomolecular pathogenesis of cholesterol gallstone disease and other diseases associated with the lipid metabolism.

Adenosine monophosphate-activated protein kinase (AMPK) activation also can inhibit the synthesis of cholesterol and fat.Recent studies have shown that activation of AMPK can also inhibit the synthesis of bile acid and promote bile salt export pump' s generation.All of those illustrated that AMPK played an important role in regulating bile acid metabolism.This article will summarize the AMPK activation pathway,bile acid metabolism,AMPK activity in bile acid synthesis and transfer,and so on.

Objective Giardia lamblia is an important pathogen of zoonosis giardiasis , it poses a potential threat to the quality of SPF (specific pathogen-free) laboratory animals cannot be ignored.The aim of this study is to establish the method of rapid diagnosis of Giardia lamblia, and analyze the test results of 516 batches form 17 manufactures.Methods Direct microscopy, Giemsa-fast staining and multiplex polymerase chain reaction (multiplex PCR) were applied to detect Giardia lamblia.Results Numerous of Giardia lamblia trophozoites and cysts were detected in SPF laboratory animals by using direct microscopy and Giemsa-fast staining, and multiplex PCR were performed to identify 18S rDNA,β-giardin, TPI and GDH genes of DNA extracted from these trophozoites and cysts identified Giardia lamblia.Direct microscopy, Giemsa-fast staining, and multiplex PCR methods can be used to detect Giardia lamblia.Of the 2562 SPF laboratory animals studied, 22.9%(586/2562) were positive for Giardia lamblia.Conclusions Direct microscopy , Giemsa-fast staining , and multiplex PCR were effective techniques with high sensitivity and specificity for rapid diagnosis of Giardia lambliain.It is not satisfactory that the results of Giardia lamblia examination in 516 batches form 17 manufactures failed to meet the requirements 100%.

Objective To diagnosis tumor transplanted nude mice Strongyloidiasis .Methods Postmortem microscopic examination of the tumor transplanted nude mice detected Strongyloides stercoralis for morphological identification and double polymerase chain reaction ( PCR ) assay for molecular diagnosis of Strongyloides stercoralis infection in tumor transplanted nude mice .Results Presence of numerous S .stercoralis in autopsy in tumor transplanted nude mice samples preliminary determined the diagnosis of strongyloidiasis .Confirmed diagnosis of Strongyloides stercoralis infection by double PCR detection of specific DNA in tumor transplanted nude mice samples .Conclusion The most important clue to prevent such serious consequences is early diagnosis .Tumor transplanted recipients and donors should be screened for parasitic infections including strongyloidiasis .To the authors ’ knowledge , this study is the first extensive report on diagnosis tumor transplanted nude mice Strongyloidiasis .

AIM:To investigate the effects of leptin on the expression of bile salt export pump ( BSEP) and signaling pathway in human hepatocellular carcinoma cell line HepG2.METHODS: HepG2 cells were cultured in vitro. Leptin at concentrations of 10 -8 , 10 -7 and 10 -6 mol/L was used as a stimulating factor.The protein levels of adenosine monophosphate-activated protein kinase alpha subunit (AMPKa), phosphorylated AMPKa (p-AMPKa) and BSEP in the HepG2 cells at 24 h, 48 h and 72 h were detected by Western blotting.The optimal culture time and leptin concentration were selected, and compound C at concentration of 10 μmol/L was added to this group.The protein expression of BSEP was detected by Western blotting.RESULTS:Intervention of HepG2 cells with leptin for 72 h increased the protein ex-pression of AMPKa gradually in a concentration-dependent manner, and leptin at concentration of 10 -6 mol/L induced the strongest AMPKa expression ( P<0.01 ) .Intervention of HepG2 cells with leptin for 24 h increased the phosphorylation level of AMPKa gradually in a dose-dependent manner (P<0.01).The effect of leptin on the increase in the protein ex-pression of p-AMPKa was also in a time-dependent manner ( P<0.01) .After intervention with different concentrations of leptin for 24 h, the protein expression of BSEP in the HepG2 cells was gradually increased by the stimulation of leptin in a concentration-and time-dependent manner (P<0.01).Compared with NC group, the protein expression of BSEP in 10 -6 mol/L leptin group and 10 -6 mol/L leptin+10μmol/L compound C group was increased at 72 h (P<0.01), and that in 10-6 mol/L leptin+10 μmol/L compound C group was lower than that in 10 -6 mol/L leptin group at 72 h ( P<0.01 ) . CONCLUSION:Leptin promotes the protein expression of BSEP in HepG2 cells by leptin-AMPK-BSEP signaling path-way.Leptin promotes the increases in AMPKa protein and the level of phosphorylation of AMPKa in HepG2 cells.

Objective To develop a multiplex polymerase chain reaction ( mPCR) assay for detection of four pathogenic dermatophytes [Trichophyton mentagrophytes (Tm), Microsporum gypseum (Mg), Microsporum canis (Mc), and Arthroderma simii ( As) ] in laboratory animals, which could be used rapidly and simultaneously for direct detection of those four pathogens.Methods We designed 5 specific primers according to 18S-28S rRNA sequences of the four pathogenic dermatophytes reported in Genbank. The four mPCR assays were established through optimizing the concentration of primers, dNTP, TaqDNA polymerase and the annealing temperature.After verifying the specificity and sensibility, this method was used to detect 15 hair samples with artificial infection and 260 samples taken from laboratory animals.Results This mPCR technique can distinguish the four dermatophytes by producing 192 bp( Tm) ,460 bp( Mg) , 290 bp( Mc) and 602 bp( As) fragments.The sensibility for detection of the four dermatophytes was 5.9 pg/μL, 6.6 pg/μL, 9.5 pg/μL and 5.1 pg/μL, respectively.The results of 15 artificial infection samples were accurate, and the results of 260 hairs samples were negative for the four fungi.Conclusions Our results suggest that the mPCR assay developed in this study can efficiently detect the four dermatophytes, is a useful and rapid technique for rapid detection of the pathogenic dermatophytes in laboratory animals.

Objective To investigate the detection capacity of esterase-3 ( Es-3) in the laboratory animals monito-ring laboratories in China, and to improve the quality management of laboratories.Methods We prepared the test sam-ples according to the criteria of China National Accreditation Service for Conformity Assessment(CNAS), all the samples were certificated by homogeneity test and stability test.Then, samples with random numbers and standard operation instruc-tion were distributed to the participant laboratories.The laboratories should submit their reports before the deadline expires. When the results are the same as the standard results, the laboratories will receive excellent remark; when the results are the same as the standard results except the hybridization type, the laboratories will receive satisfactory remark;otherwise, it will receive unsatisfactory remark.If a laboratory did not submit report, the laboratory will also receive unsatisfactory re-mark.Results Ten laboratories participated in the program, and no laboratory received excellent remark.Nine laboratories (90.0%of enrolled laboratories) had satisfactory results, while one laboratory (10.0%of enrolled laboratories) had un-satisfactory results.Conclusions The nationwide overall detection level of laboratories in Es-3 is relatively high.Howev-er, some details should be noticed and several laboratories should improve their detecting ability.

0.05). Cross-reactivity was seen in 1 case of clonorchiasis sinensis by both the methods. Conclusion Microwave ELISA has the advantages of high sensitivity, specificity and rapidity.

Objective To test the potency of hepatitis B vaccine in China. Methods Two inbred strains(DBA/1 and BALB/c) and two NIH closes-colonies of mice were typed in the H-2 region by microcy-totoxicity method and PCR. Groups of mice of the tested strains were immunized with the same hepatitis B vaccine, the titre of anti-HBsAg antibody was analyzed by microplate, and the ED50 was then estimated by Karder method for each strain. Results Significant differences were found between potency estimates de-rived from assays using different strains of mice. Conclusion It is likely that the variation of immune re-sponse to hepatitis B vaccine in mice is correlative with the H-2 haplotype. In some special case, the bet-erozygosity in H-2 region found in NIH stock could influence the accuracy in such testing even a reference preparation of hepatitis B vaccine was used. Base on our experiment, to select an appropriate NIH stocks with the H-2q haplotype for potency testing of hepatitis B vaccine in China.