Wu Jin-Ya, Zhou Xiao-Hong, Chen Xiao-Guang
Department of Parasitology, Southern Medical University, Guangzhou 510515, China.
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2005 Apr 30;23(2):110-3.
To express P38 of Schistosoma japonicum in Escherichia coli BL21 and develop the monoclonal antibodies (McAb) against rSjP38.
The recombinant plasmid pET32(a)-P38 was transformed into E. coli BL21 (DE3). 1 mmol/L IPTG (isopropyl-beta D-thiogalactopyranoside) was used to induce the expression of the recombinant rSjP38. The rSjP38 was soluble in supernatant after sonication and further purified by His-Ni chromatography. BALB/c mice were immunized with the purified rSjP38 and hybridomas were generated with traditional technique. McAbs were screened by ELISA with limited dilution. The subtype and specificity of McAb were identified by kit and Western blot respectively.
Eight hybridoma cell lines secreting monoclonal antibodies to rSjP38 were obtained. The subtype of all the 8 McAbs are IgG1. Western blotting showed that the 8 McAbs reacted strongly and specifically with native antigen (P38) of Schistosoma japonicum.
Eight hybridoma cell lines secreting highly specific McAbs against P38 have been established.
在大肠杆菌BL21中表达日本血吸虫P38，并制备抗重组日本血吸虫P38（rSjP38）的单克隆抗体（McAb）。
将重组质粒pET32(a)-P38转化至大肠杆菌BL21(DE3)。用1 mmol/L异丙基-β-D-硫代半乳糖苷（IPTG）诱导重组rSjP38表达。超声破碎后，rSjP38可溶在上清中，再经His-Ni柱层析进一步纯化。用纯化的rSjP38免疫BALB/c小鼠，采用传统技术制备杂交瘤。通过有限稀释法用ELISA筛选单克隆抗体。分别用试剂盒和Western blot鉴定单克隆抗体的亚型和特异性。
获得8株分泌抗rSjP38单克隆抗体的杂交瘤细胞株。8株单克隆抗体的亚型均为IgG1。Western blotting显示，这8株单克隆抗体与日本血吸虫天然抗原（P38）反应强烈且特异。
已建立8株分泌针对P38的高特异性单克隆抗体的杂交瘤细胞株。
