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Effect of Storing Time of Venous Blood Samples on the Differential Counts of White Blood Cells by Sysmex XE-2100 Hematology Analyzer

Weizhen FANG ; Mei CHEN ; Yuru FU ; Changzhen XU

Journal of Tropical Medicine.2005;5(6):786-789,799.

Objective To evaluate the influence of the storing time of venous blood samples on the differential count of WBC by Sysmex XE-2100 hematology analyzer. Methods At room temperature, the precision of the differential count of WBC by Sysmex XE-2100 hematology analyzer were tested. 38 samples were taken the differential count of WBC by Sysmex XE-2100 hematology analyzer after stored for 0, 2, 4, 8, 24 and 48 h. Differential count of WBC was also taken under microscope for comparison. Results The precision of differential count of WBC by Sysmex XE-2100for all the samples was in the allowable range. The correlation coefficient of the differential count of WBC by two methods for neutrophils, lymphocytes, monocytes, eosinophils and basophils were 0.9859, 0.9775, 0.8053, 0.8695and 0.5243 (P<0.01). There was insignificant difference in the test at 8h, very significant difference of MONO and EOS at 48 h. Differences of EOS, MONO between two methods were significant increased at 8 h. Conclusion At room temperature, the differential count of WBC of venous blood samples by Sysmex XE-2100 hematology analyzer should finish within 8 h.

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Evaluation on the Identification and Counting of Nucleated Red Blood Cells by Sysmex XE-2100 Hematology Analyzer

Mei CHEN ; Weizhen FANG ; Yuru FU ; Qiongzhu LIN ; Changzhen XU

Journal of Tropical Medicine.2005;5(1):45-48,25.

Objective To evaluate the identification and counting effeciency of nucleated red blood cells by Sysmex XE-2100 Hematology Analyzer. Methods Accurancy: nucleated red blood cells were counted from 38 specimens by Sysmex XE-2100 Hematology Analyzer and compared with the number counted under microscopy. Precision: 3 specimens with different values were counted for the nucleated red blood cells 10 times by Sysmex XE-2100 Hematology Analyzer and compared with the number counted under microscopy. The CV(% ) value was estimated.Results There was insignificant difference between the results obtained from Sysmex XE-2100 Hematology Analyzer and those under microscopy. In the T-test, P >0.05, r=0.9893(P< 0.01).CV(% ) were 8.1% and 15.8% . It means that the Sysmex XE-2100 is more precise in analyzing the nucleated red blood cells than that under microscopy. Conclusion The nucleated red blood cells count by Sysmex XE-2100 is accurate and fast to obtain clinical data.

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Molecular Cloning and Characterization of Two Hommologues of Yeast Sir2 Gene in the Parasitic Protist Trichomonas vaginalis

Yongmei SHI ; Yucai FU ; Mingyan XU ; Xiaoyuan XU ; Jinjie XU ; Tang PETRUS

Journal of Tropical Medicine.2007;7(4):297-302,306.

Objective To screen cell growth and senescence-related genes of the parasitic pmtist Trichomonas vaginalis,we launched an EST program and isolated two cDNA clones from a T.vaginalis cDNA library,which showed high homology in deduced amino acid sequences to yeast Sir2 and designated as TvSir2 and TvSir2-like.Method The cDNA sequence of TvSIR2 had a length of 1034 base pairs (bp) with an open reading frame of 915 bp,and TvSIR2-like,1214 bp with an open reading frame of 1116 bp.Result The two deduced amino acid sequences shared all the three conserved cole domains with yeast Sir2 and its homologues,suggesting that the two clones were Sir2 homologues. A cDNA fragment from each cDNA clone was subvloned into the expression vector pET-41a.The expression of the fusion proteins in E.coli BL21 stains was induced by isopropylthio-β-D-galactoside (IPTG).Two anti-sera were prepared by immunizing two guinea pigs with the purified fusion proteins, Western-blot analysis demonstrated that each anti-serum reacted with the corresponding recombinant protein and detected a clear band (TvSir2,34 000 Mr;TvSir2-like,42 000 Mr)in protein extracts of the protist.Immunofluolescence techniques showed that TvSir2 and TvSir2-like proteins were both localized in the legions of perinuelear (ER) and Golgi complex.Conclusion Our data suggest that TvSir2 and TvSir2-like were two members of Sir2 family.Their biological functions in the protist would be further studied.

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Determination of Circulating Antigen in Neurocysticercosis Patients by Monoclonal Antibody

Zengzhu GUO ; Minjun HUANG ; Yijun AN ; Song HUANG ; Hongjie JIANG

Journal of Tropical Medicine.2005;5(2):113-115.

Objective To detect circulating antigen (Cag) for diagnosing neurocysticercosis. Method ELISA was performed with monoclonal antibody 4B6 against the cyst fluid antigen of Cyticercus cellulosae for detecting Cag in serum and/or cerebrospinal fluid (CSF) of patients with neurocysticercosis or with other diseases. Results In the group of 82 cases of neurocysticercosis, the positive rate of serum Cag was 79.2% (65/82) and the positive rate of CSF Cag was 100% (26/26). After chemotherapy for 20 cases with positive serum Cag, the titer of serum Cag in 17 cases dropped to zero(85% ). Cag could not be detected in specimens from patients with other diseases. Conclusion These results indicate that the determination of Cag, especially of the CSF Cag, is useful for the diagnosis of neurocysticercosis and the drop in serum Cag is a good parameter for the evaluation of the effectiveness of chemotherapy.

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Animal Host of Schistosoma japonicum and Transmission of Schistosomiasis in Poyang Lake Region

Dandan LIN ; Yueming LIU ; Fei HU ; Shaoji ZHANG

Journal of Tropical Medicine.2003;3(4):383-387.

The article clarified, according to the recent research results which have been done in Poyang Lake region of Jiangxi province, bovines are the most important animal hosts of Schistosoma japonicum and infected bovines are main infectious sources of Schistosomiasis in Poyang Lake region. Spring (March to June) is the " cross transmission stage" between definitive host and intermediate host, that is, spring is not only the susceptible season of infection for livestock and human, but also is the susceptible season of infection for snails. In flood season (July to August), the transmission of Schistosomiasis between livestock and snails belong to " low level phase" . Autumn (September to Octber) is the second seasonal peak of cercariae and is the susceptible season of infection for livestock and human. Winter(November to Feburary in next year) is the non- susceptible season of infection. Most of the susceptible zones of infection for livestock, human and snails are the marshlands near the endemic villages.

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Primary Study of the Structure and Antigenic Epitopes of Antigen Pf332

Zhixin SHAN ; Xinbing YU ; Jin XU ; Zhongdao WU ; Xuerong LI ; Guowu BIAN ; Changling MA ; Yan LI ; Shouyi CHEN ; Xuchu HU

Journal of Tropical Medicine.2002;2(3):219-224.

Objective To understand the primary structure and potential antigenic epitopes of antigen Pf332(Ag332) of P.falciparum iso late FCC1/HN.Methods　Based on the published Pf332 gene sequence , nine pairs of primers were designed for the PCR amplification of the Pf332 gen e fragments from genomic DNA of P.falciparum isolate FCC1/HN. The amplified gene fragments were subcloned into pMD-18T vectors and sequenced. The sequences were aligned using DNAstar software to obtain the full-length sequence of the gene Pf332. The primary structure and sequence homology of Ag332 were analyzed by SAPS, Tmpred, SingalP and Blastn programs. Three fragments, R0, R1 and R2, cor responding to nt#9595-10083, nt#10339-10767 and nt#10855-11247 of Pf332 gene were subcloned into the eukaryotic expression vector pcDNA3-S separately. The Balb/c mice were immunized with pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3- S-R2 separately, and the expressions of the recombinant proteins were detected by immunohistochemistry assay. The protective immune responses elicited by DNA I mmunization were analyzed by ELISA and parasite growth inhibition tests in vitro .Results　Nine Pf332 gene fragments were specifically amplif ied, subcloned into pMD-18T vectors and sequenced. Pf332 gene of the P.falci parum isolate FCC1/HN was 16,377 bp in length, encoding a protein of 5,458 ami no acids, about 615.28kDa. The Ag332 contains 17 regions of highly degenerated Glu-rich repeats, with 30.18% Glu in total amino acids of Ag332. Ag332 of P.falciparum isolate FCC1/HN and 3D7 exhibited 94.55 % homology in amino acid residues. The results of immunohischemistry assay showed that R0, R1 and R2 were expressed in mice muscle tissue. The amount of IgG antibody of the groups immu nized with pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3-S-R2 were higher than those of blank and pcDNA3 groups (P＜0.05). The result of parasite growth inhibition test showed that the immunized sera at 1∶5 dilution of groups of pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3-S-R2 had an incomplete inhibitor y effect on P.falciparum growth. Conclusion　The antigen Pf332 is an large protein containing highly degenerated Glu-rich repeats. Pf332 gene fragments, R1 and R2 encoding potent antigenic epitope repeats.

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The genetic immunization in mice with three constructs of Toxoplasma gondii SAG1 gene

Xiaoguang CHEN ; Xiaohong ZHOU ; Ya GONG ; Peiliang YANG ; Shuman SHEN ; Mingchiu FONG ; Zhaorong LUN

Journal of Tropical Medicine.2001;1(1):3-9.

Three forms of the major surface antigen (SAG1)of Toxoplasma gondii, that is the membrane form, the secrete form and the intracellular form, were constructed and used to immunize BALB/c mice. The humoral response in the mice immunized with the membrane form and the secrete form of SAG1 appeared earlier and stronger than those mice immunized with the intracellular form. Result from the challenging infection demonstrated that the protection in the mice immunized with the membrane and the secrete forms was also stronger than in the mice immunized with the intracellular form. We suggest that the immune efficiency of the three forms of SAG1 in the mouse model is different.

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Human Cytomegalovirus UL143 Gene Polymorphisms in Low-Passage Clinical Isolates in Guangzhou

Bo WANG ; Yueqing LI ; Ning YE ; Jingjing HU ; Haihao SU ; Zhenyu HE ; Chuanjun TIAN ; Chunqing ZHANG ; Tianhong ZHOU

Journal of Tropical Medicine.2008;8(4):327-331.

Objective To investigate the polymorphism of human cytomegalovius UL143 gene of low passage clinical isolates in Guangzhou,China.Method PCR was performed to amplify the entire HCMV ULl43 gene region of 3 clinical isolates,which had been proven by multiplex PCR.The amplification products were cloned into pMD18-T-Vector and subjected to sequencing.The result of DNA sequences were analyzed together with the one of published homologous sequences in GenBank from 14 clinical isolates.Result There were several stop codons in UL143 gene due to a base deletion in open reading frame (ORF) of D3 isolate,which could lead to produce non-functional protein.UL143 ORF of Toledo isolate consisted of 279 nueleotides,encoding a protein with 92 amino acids.UL143 ORFs of other isolates consisted of 252 nueleotides,encoding a protein with 83 amino acids.The DNA sequences were quite conserved and all the variations were base substitution.The amino acid sequences of different isolates were highly conserved.with variation of 1.2%-2.4%.There were no additional or deleted sites of post translational modification of UL143 protein in all clinical isolates except Toledo isolate.There were some differences in the secondary structure among different isolates.The isoelectric point of UL143 protein of all clinical isolates except Toledo isolate was 8.75.Conclusion All DNA and deduced amino acid sequences of UL143 gene shared great similarity among HCMV clinical strains regardless of their polymorphism.

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Expression of Human Connexin26 in COS-7 Cells

Cong XU ; Guowei LI ; Zeyu LIU ; Jiawei LIAO ; Panpan XUE ; Binyan TAN ; Jiahai LU

Journal of Tropical Medicine.2007;7(5):401-407.

Objective To construct the eukaryotic expression plasmid for the expression of human Connexin26 in COS-7 cells.Methods Total RNA was isolated from human peripheral blood lymphocytes and used as template for the PCR cloning of the human Connexin26 gene.The human Cx26 cDNA containing the 678 bp whole coding region of the human Connexin26 gene was amplified by PCR using specific primers and cloned into the pCI-neo vector to construct the recombinant eukaryotic expression plasmid,pCI-Cx26.The recombinant plasmid was identified by restriction endonuclease digestion,and transfected into COS-7 cells by liposome.The expression of Cx26 mRNA and the protein were analyzed by RT-PCR and SDS-PAGE,respectively.Results Restriction endonuclease digestion analysis verified successful construction of the recombinant plasmid,pCI-Cx26.The expression of Cx26 mRNA and protein in the transfected COS-7 cells were detected by RT-PCR and SDS-PAGE,respectively.Conclusion The eukaryotic expression plasmid for human Cx26 has been constructed successfully with the capability of expression in COS-7 cells.

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Classification of A Medically Important Group of Gamasid Mites by Numerical Taxonomy in Yunnan, China

Lipu LUO ; Xianguo GUO

Journal of Tropical Medicine.2007;7(1):7-10.

Objective To determine the taxonomic position of medically important gamasid mites and to compare numerical taxonomy with the traditional systematics in the classification of gamasid mites. Methods Based on hierarchical cluster analysis, numerical taxonomy was applied to study fifty-seven species of medically important gamasid mites. Results The results of cluster analysis based on squared euclidean distance showed that Hirstionyssus Fonseca and Echinonyssus Hirst should be separated from Laelapidae Berlese and formed an independent family. The taxonomic position of the remaining species remains the same as those in the traditional systematics. Conclusion Numerical taxonomy can objectively reflect the taxonomic position of the medically important gamasid mites. The result of classification by numerical taxonomy is consistent with traditional systematics for gamasid mites.

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