In the present study , the express library of monoclonal anti - sp18 scfv ( single chain fragment variable ) gene is constructed and selected for further study of sp18 antigen on mammalian fertilization and embryogenesis . total rna were firstly isolated from these growing hybridoma cells which secretes monoclonal anti - sp18 antibodies . after obtained using rpas system , vh and vl genes were used to assemble scfv gene fragment with a linker primer 应用重组噬菌体抗体库技术，从分泌小鼠抗牛精子sp18抗体的杂交瘤细胞系中分离总rna ，克隆抗体重链和轻链可变区基因，加入连接肽引物( linkerprimer )组装成单链抗体scfv ( singlechainfragmentvariable )基因并用rs引物进行扩增， sfi 、 not酶切，回收后与pcantab5e载体相连，转化e . colitg1宿主菌，构建单链抗体文库。
Four mcabs were developed and can detect the meq gene expression in the cef infected with oncogenic mdvs and naturally occurred md tumor cells by fa and immunohistochemistry technique respectively , but ca n ' t detect that in the cef and normal chicken liver tissue respectively infected with non - virulent vaccine virus cvi988 / rispens . the results demonstrated that meq is highly expressed in t 获得4株稳定产生抗meq蛋白mcab的杂交瘤细胞，其中3g12e6单抗通过fa和免疫酶组化技术能够检测到mdv致瘤株感染的cef及自然md肿瘤细胞中表达的me叮基因产物，而在非致瘤株cvi988 / rispens感染的cef和免疫的正常鸡的组织中则未检测到。
2 - e4 - a and 82 - 6 are hybridized during their log growing time , and the hybrid - hybridomas are cloned for 3 times and produce 6 hybrid - hybridoma cells . the chromatosome of hybrid - hybridoma 3 - hu and hybridoma 2 - e4 - a and s2 - b are counted , and the antibody of ascites fluid or culture supernatant of 3 - hn is prepared . the positive clones are detected by three methods at the same time : rbc agglutination for monospecific anti - human rbc type a antibody , indirect elisa for anti - p24 antibody , and rbc solid - phase adherence for bispecific antibody 选其中一株3 - h _ ( 11 )做杂交-杂交瘤细胞染色体计数，同时计数两母株2 - e _ 4 - a和s _ 2 - b的染色体数：制备腹水型和上清型抗体，用三种方法同时检测其中的双特异性抗体、单特异性抗人红细胞抗体和抗p24抗体，即红细胞固相吸附法测双特异中文摘要性抗体，红细胞凝集试验测单特异性抗人a型红细胞抗体，间接elisa法测抗p24抗体；用腹水型抗体做耐热性及耐冻融实验。
Culture of mg7 hybridoma cells and detection of antigen - binding affinity of mg7 mab by elisa 2 . construction and identification of mg7 recombinant phage antibody library mrna was isolated from cultured mg7 hybridoma cells and converted into cdna ; the variable fragments of heavy and light chain were separately amplified and assembled into scfvs with a specially constructed dna linker by pcr . the scfvs dma was ligated into the phagmid vector pcantabse and the ligated sample was transfered into competent e . co / / tg1 to generate a bacterial form of mg7recombinant phage antibody library ; volume and recombinant ratio of the library were evaluated by means of bacterial colony counts and restriction analysis ( ecor i and hind iii ) Mg _ 7重组噬菌体抗体库的构建及鉴定从培养的mg _ 7杂交瘤细胞中提取并分离mrna ，反转录成cdna ；利用pcr分别扩增mg _ 7单抗的重链及轻链可变区基因，并通过? dna连接子将二者连接起来形成mg _ 7单链抗体基因；将mg _ 7单链抗体基因插入pcantab5e ；将连接产物转化感受态tg1大肠杆菌，制备细菌形式的mg _ 7重组噬菌体抗体库；通过菌落计数和限制性酶切分析( ecor和hind )评估mg _ 7重组噬菌体抗体库的容量和重组率。
