Sripa Jittiyawadee, Brindley Paul J, Sripa Banchob, Loukas Alex, Kaewkes Sasithorn, Laha Thewarach
Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
Parasitol Int. 2012 Mar;61(1):191-5. doi: 10.1016/j.parint.2011.05.009. Epub 2011 Jun 17.
A cathepsin B-like cysteine protease belonging to family C1 is abundantly expressed in the transcriptome and proteome of the carcinogenic liver fluke of humans, Opisthorchis viverrini. This enzyme is present in excretory/secretory (ES) products released by parasites cultured in vitro. This study evaluated the performance of recombinant O. viverrini cathepsin B1 (rOv-CB-1) as an antigen for immunodiagnosis of opisthorchiasis. The full length Ov-CB-1 cDNA was cloned and recombinant protein was produced in catalytically active form in Pichia pastoris. The recombinant Ov-CB-1 (rOv-CB-1) was affinity purified via nickel-NTA chromatography and tested in enzyme-linked immunosorbent assays (ELISA) with human sera from an opisthorchiasis endemic area. Sera from egg-positive O. viverrini infections produced a strong IgG antibody response to rOv-CB-1 both in ELISA and immunoblot analysis. The sensitivity and specificity of the ELISA test was 67% and 81%, respectively. These findings support the feasibility of using recombinant Ov-CB-1 in ELISA for the serodiagnosis of human opisthorchiasis.
一种属于C1家族的组织蛋白酶B样半胱氨酸蛋白酶在人类致癌肝吸虫——华支睾吸虫的转录组和蛋白质组中大量表达。这种酶存在于体外培养寄生虫释放的排泄/分泌（ES）产物中。本研究评估了重组华支睾吸虫组织蛋白酶B1（rOv-CB-1）作为华支睾吸虫病免疫诊断抗原的性能。克隆了全长Ov-CB-1 cDNA，并在毕赤酵母中产生了具有催化活性形式的重组蛋白。重组Ov-CB-1（rOv-CB-1）通过镍-亚氨基三乙酸（NTA）色谱进行亲和纯化，并在酶联免疫吸附测定（ELISA）中用来自华支睾吸虫病流行地区的人血清进行检测。在ELISA和免疫印迹分析中，来自华支睾吸虫卵阳性感染的血清对rOv-CB-1产生了强烈的IgG抗体反应。ELISA检测的敏感性和特异性分别为67%和81%。这些发现支持了在ELISA中使用重组Ov-CB-1进行人类华支睾吸虫病血清诊断的可行性。
