Xi-Bin Xiao

Sun Yat-Sen University, Guangzhou, Guangdong Sheng, China

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Publications (6)0 Total impact

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    ABSTRACT: To study the renaturation, purification and binding activity of scFv of anti-nasopharyngeal carcinoma monoclonal antibody(mAb) BAC(5) expressed as inclusion body in E.coli. The E.coli BL21(DE3) transformed with the pET 22b-scFv was cultured and pulvereged by ultrasonic cell disintegrator. The collected inclusion bodies were denatured with 8 mol/L urea and renatured by dilution refolding, step dialysis and gel filtration chromatography. Binding activity of renatured BAC(5)-scFv was determined by immunohistochemical staining and Western blot. BAC(5)-scFv purified though Ni-NTA His Bind chromatographic clomn showed high purity. The highest proteins recovery rate was obtained through gel filtration chromatography. It was proved by Western blot and immunocytochemical staining that the renatured BAC(5)-scFv protein could specifically bind to CNE2 cells. BAC(5)-scFv expressed as inclusion body retained good activity after being dissolved, purified and renatured, which paves the way for preparing large amount of BAC(5)-scFv to be used for the study of radioimmunoimaging and therapy of nasopharyngeal carcinoma.
    Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology 02/2006; 22(1):67-70.
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    ABSTRACT: Nasopharyngeal carcinoma (NPC) patients commonly have neck regional lymph node metastases. However, patients with NPC of stage T3-4N0 have no lymph node metastases, even the tumors grow over the nasopharyngeal cavities. This study was to explore immune factors related to such a situation by detecting gene expression profile of immune cells from patients with NPC of stage T3-4N0. Peripheral blood lymphocytes were isolated from 28 patients with NPC of stage T1-2N2-3, 28 patients with NPC of stage T3-4N0 before treatment, and 56 healthy donors. mRNA of the isolated lymphocytes was extracted, and reverse-transcribed into cDNA probes labeled with Cy5 or Cy3 fluorescent dyes. The Cy5-cDNA probes of T1-2N2-3 group and T3-4N0 group were separately mixed with Cy3-cDNA probes of healthy group, and hybridized with 480 dots of immune genechips. Genes were considered to be differently expressed in NPC patients if the Cy5/Cy3 was >2.0 (up-regulated), or <0.5 (down-regulated). The genes with Cy5/Cy3 of <0.25 or >4.0 were amplified by real time quantitative reverse transcription-polymerase chain reaction (RT-PCR) with RNA templates of T1-2N2-3 group, and T3-4N0 group, respectively. Of 157 genes showed different expressions, 106 down-regulated, 51 up-regulated. Real time quantitative RT-PCR revealed that among the 4 genes with Cy5/Cy3 of >4.0, and the 3 genes with Cy5/Cy3 of <0.25, expressions of the genes encoding CD86, CD69, NFKB were higher in sample from T3-4N0 group than in sample from T1-2N2-3 group. More genes are down-regulated in immune cells from patients with NPC of stage T1-2N2-3 or T3-4N0 than those from healthy donors. Whether the different expressions of genes encoding CD86, CD69, and NFKB in immune cells of patients with NPC of stage T1-2N2-3 and stage T3-4N0 related to neck regional lymph node metastasis is worthy of further study.
    Ai zheng = Aizheng = Chinese journal of cancer 02/2005; 24(2):135-40.
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    ABSTRACT: Epstein-Barr virus (EBV) is closely related to nasopharyngeal carcinoma (NPC). Many kinds of methods can be used to examine antibodies in NPC patients sera. This study was to screen the dominant epitopes from random peptide libraries (RPLs) displayed on phage using the EBV-related antibodies purified from the sera of NPC patients, and find new antigens at the epitope level. The EBV-related antibodies were eluted from the sera of NPC patients using B95-8 cell EBV proteins as antigen, and the phages from 12-mer RPLs were elutriated for 3 rounds with the antibodies. The positive clones were gained by sandwich ELISA, and competitive inhibition assay from the third elution. The positive clones were sequenced, and the peptide coded by the inserted DNA were blasted with the antigen region of EBV proteins. Sixty-four phage clones were randomly picked up from the third eluate,25 positive clones were picked up using sandwich ELISA assay, the positive percentage was 39.06%. Thirteen clones were picked up for competitive ELISA assay,11 clones showed inhibitory phenomena,the inhibitory rates were between 18.09% and 65.94%. Five positive clones with high absorbency value, and high inhibitory rates were selected out, the sequences of peptides displayed on these clones were -A-T-S-H-L-H-V-R-L-P-W-T- (d15, and d18), -G-S-T-H-K-H-N-H-F-N-K-T- (d19), -K-P-I-H-E-H-P-H-R-F-K-S- (e8), -H-T-H-K-I-K-I-P-L-P-I-Q- (e23). These peptide sequences showed similarity with the amino acid sequences located in antigen regions of EBV integral membrane protein (d15, and d18), EBV thymidine kinase (d19), and EBV major capsid protein (e8, and e23). EBV-related epitopes could be obtained by screening the phages from RPLs with polyclonal antibodies purified from the sera of NPC patients, which may offer new methods of antigen preparation for sera diagnosis of NPC.
    Ai zheng = Aizheng = Chinese journal of cancer 09/2004; 23(9):999-1004.
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    ABSTRACT: Combined therapy has been advocated for modern tumor treatment; the combined target therapy is a valuable research direction. Based on the previous research of nasopharyngeal carcinoma (NPC) radioimmunotherapy, this experiment was designed to develop two immunoconjugates of the monoclonal antibody BAC(5):PYM-BAC(5) and (131)I-BAC(5), and to assess the inhibition effects of their combined treatment on the NPC CNE-2 cells cultured in vitro. Dextran T40 was used as media to link PYM and BAC(5). The conjugate PYM-BAC(5) was identified by testing its immunoactivity and the inhibition to mycobacterium. BAC(5) was labeled with (131)I by Chloramin-T method. Five experimental groups were set up:(1)PYM-BAC(5) group, (2)free PYM group, (3)(131)I-BAC(5) group, (4)(131)I-mIgG group, (5)the combined target treatment group ( (131)I-BAC(5)+PYM-BAC(5)). The antitumor effects of the five groups were assessed with MTT method. The 50% inhibition doses(IC(50)) of PYM-BAC(5) group and PYM group were 46.57 microg/ml and 316.7 microg/ml, respectively. The IC(50) of (131)I-BAC(5) group and (131)I-mIgG group to CNE2 were 4.42 x 10(5) Bq/ml and >11.1 x 10(5) Bq/ml,respectively. In the combined target treatment group(PYM-BAC(5)+(131)I-BAC(5)),the IC(50) of PYM-BAC(5) was 7.01 microg/ml and of (131)I-BAC(5) was 0.54 x 10(5) Bq/ml, which much less than other separate treatment groups. The inhibition effects of the target treatment ((131)I-BAC(5) and PYM-BAC(5)) on the NPC CNE-2 cells are stronger than non-target treatment (free PYM and (131)I-BAC(5)). The combined target treatment of the two immune ((131)I-BAC(5)+PYM-BAC(5)) conjugates gets stronger inhibition effects than their separate treatment.
    Ai zheng = Aizheng = Chinese journal of cancer 08/2003; 22(8):831-5.
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    ABSTRACT: It has been reported that cell-free Epstein-Barr virus (EBV-DNA) in plasma was useful in diagnosing and monitoring nasopharyngeal carcinoma (NPC). The current study was designed to evaluate the significance of EBV-DNA in monitoring the prognosis of nasopharyngeal carcinoma and to compare with VCA/IgA and EA/IgA. EBV-DNA, VCA/IgA, and EA/IgA levels in plasma were detected in different NPC patients after radiotherapy, including 30 distant metastasis patients, 22 locoregional recurrence patients, 24 remission individuals who had been followed up more than 2 years after treatment. EBV-DNA was detected using real-time quantitative PCR system;VCA/IgA and EA/IgA were tested using regular immunofluorescence. In cohort study, the indexes were tested in different radiation periods for the 20 new cases of nasopharyngeal carcinoma. The median plasma EBV-DNA concentration was 135,100 copies/ml (interquartile range: 5,525-1,003,750) in metastasis group, 20,500 copies/ml (interquartile range: 0-58,500) in locoregional recurrence group and 0 copies/ml (interquartile range: 0-0) in continuous remission group (P< 0.05). The levels of VCA/IgA and EA/IgA had no significant difference in different groups. The high level of EBV-DNA concentration in metastasis group was more than that in locoregional recurrence group. At the level of 1,000,000 copies/ml, EBV DNA indicated distant metastasis of NPC with a specificity of 100% and a sensitivity of 27.3%; however, the sensitivity was 0 copies/ml in locoregional recurrence group. For the 20 new patients, EBV DNA concentration gradually decreased in the radiation period, 32,050 copies/ml (interquartile range: 3,880-317,750) before radiation, 0 copies/ml (interquartile range: 0-14,375) when 40 Gy radiation dose and 0 copies/ml (interquartile range: 0-2940) when the radiation finished (P< 0.05). However, the levels of VCA/IgA and EA/IgA had no significant difference. The plasma cell-free EBV-DNA is more valuable than VCA/IgA and EA/IgA for monitoring the prognosis of NPC patients.
    Ai zheng = Aizheng = Chinese journal of cancer 03/2003; 22(3):302-6.
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    ABSTRACT: The periphery T and B lymphocytes are most important immune cells in a body. Analysis for the gene expression profiles of the immune cells will contribute to the understanding of the biological essence of immune suppression. The advanced technique of DNA microarray make it possible to monitor the expression of thousands of genes simultaneously in one hybridization experiment until recently. In this paper, the authors investigated the genes differentially expression in periphery lymphocytes between the patients with Nasopharyngeal Carcinoma(NPC) and health donors to explore the features of gene expression in immune cells of NPC patients. The BioDoor10s/D chips containing 2000 spots of cDNAs were used to investigate the difference of the expression. Both the mRNAs from periphery lymphocytes of NPC patients (2 x 10(8)) and health donors(2 x 10(8)) were reversely transcribed to cDNA with the incorporation of fluorescent(cy5 and cy3) labeled dUTP to prepare the hybridization probes. After hybridization, BioDoor10s/D chips were scanned for the fluorescent intensity. The differentially expressed genes were screened. The total of 62 genes has differentially expression in NPC periphery lymphocytes. The number of over- and lower-expressed genes were 5 and 57, respectively. There were 3 genes coding for signal transductive proteins in 5 over-expressed genes. Five genes coding for signal transductive proteins were involved in 9 genes with lowest-expression (cy5/cy3 > or = 0.3). There was the consistent tendency toward lower-expressed gene in periphery lymphocytes from the patients with NPC. The disorder in signal transductive function of lymphocytes may be a factor distinguishing NPC immunity from the health one. Improvement of gene expressing profil may also be a way to enhance immune function in tumor patients.
    Ai zheng = Aizheng = Chinese journal of cancer 01/2002; 21(1):21-4.