The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lymphoma has led to new treatment options for this group of patients. Production ofmonoclonal antibodies by t...The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lymphoma has led to new treatment options for this group of patients. Production ofmonoclonal antibodies by the traditional hybridoma technology is costly, and the resulting murine antibodies often have the disadvantage of triggering human anti-mouse antibody （HAMA） response. Therefore recombinant Fab antibodies generated by the phage display technology can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cells of BALB/c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the κ light chains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successively inserted into the phagemid vector pComb3H-SS to construct an immunized Fab phage display library. The diversity of the constructed library was approximately 1.94× 10^7. Following five rounds of biopanning, soluble Fab antibodies were produced from positive clones identified by ELISA. From eight positive clones, FabC06, FabC21, FabC43 and FabC59 were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains （VH） and variable light domains （VL） were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murine germline genes respectively. Furthermore, reactivity with membrane proteins of the B cell lymphoma was demonstrated by immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for further study of B cell lymphoma and could also contribute to the improvement of disease therapy.展开更多
目的:探讨健脾祛湿方外敷治疗小儿脾虚湿蕴型湿疹的临床疗效,且对患儿免疫球蛋白及外周血γ干扰素(interferon-γ,IFN-γ)、重组人B细胞淋巴瘤蛋白(recombinant human B cell lymphoma protein,Bcl-6)水平表达的影响。方法:140例脾虚湿...目的:探讨健脾祛湿方外敷治疗小儿脾虚湿蕴型湿疹的临床疗效,且对患儿免疫球蛋白及外周血γ干扰素(interferon-γ,IFN-γ)、重组人B细胞淋巴瘤蛋白(recombinant human B cell lymphoma protein,Bcl-6)水平表达的影响。方法:140例脾虚湿蕴型湿疹患儿按随机数字表法分为两组,均为67例。对照组仅予以氯雷他定颗粒治疗,体质量≤30 kg的患儿每次5 mg,体质量30 kg以上的患儿,每次10 mg,饭后温水溶解服用,每天1次。研究组采用健脾祛湿方外敷治疗。比较两组皮损面积、瘙痒程度、中医证候评分、皮损状态、临床疗效、免疫指标、IFN-γ、Bcl-6水平变化情况。结果:两组治疗前皮损面积、瘙痒程度比较,差异无统计学意义(P>0.05),治疗后对照组皮损面积大于研究组,瘙痒程度高于研究组(P<0.05)。治疗前两组证候评分、皮损状态比较,差异无统计学意义(P>0.05),治疗后对照组证候评分、皮损状态均优于研究组(P<0.05),对照组有效率低于研究组(P<0.05)。治疗前两组免疫球蛋白M(immunoglobulin M,IgM)、免疫球蛋白G(immunoglobulin G,IgG)水平比较,差异无统计学意义(P>0.05),治疗后对照组IgM、IgG水平均高于研究组(P<0.05)。治疗前两组IFN-γ、Bcl-6水平比较,差异无统计学意义(P>0.05),治疗后对照组IFN-γ水平低于研究组,Bcl-6水平高于研究组(P<0.05)。结论:健脾祛湿方外敷治疗小儿脾虚湿蕴型湿疹效果显著。展开更多
基金This work was supported by grants from the National Natural Science Foundation of China(No.30400111)the Natural Science Foundation of Jiangsu Province(No.BK2004041).
文摘The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lymphoma has led to new treatment options for this group of patients. Production ofmonoclonal antibodies by the traditional hybridoma technology is costly, and the resulting murine antibodies often have the disadvantage of triggering human anti-mouse antibody （HAMA） response. Therefore recombinant Fab antibodies generated by the phage display technology can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cells of BALB/c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the κ light chains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successively inserted into the phagemid vector pComb3H-SS to construct an immunized Fab phage display library. The diversity of the constructed library was approximately 1.94× 10^7. Following five rounds of biopanning, soluble Fab antibodies were produced from positive clones identified by ELISA. From eight positive clones, FabC06, FabC21, FabC43 and FabC59 were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains （VH） and variable light domains （VL） were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murine germline genes respectively. Furthermore, reactivity with membrane proteins of the B cell lymphoma was demonstrated by immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for further study of B cell lymphoma and could also contribute to the improvement of disease therapy.
