National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
Exp Parasitol. 2011 Jan;127(1):287-93. doi: 10.1016/j.exppara.2010.06.018. Epub 2010 Jun 25.
A novel gene, BmP94, encoding 94-kDa protein of Babesia microti was identified by immunoscreening of the cDNA expression library. The full-length of BmP94 was expressed in Escherichia coli (rBmP94), which resulted in insoluble form with low yield, and the truncated hydrophilic C-terminus region of the gene was expressed as a soluble protein (rBmP94/CT) with improved productivity. Antiserum raised against rBmP94/CT recognized the 94-kDa native protein in the parasite extract by Western blot analysis. Next, an ELISA using rBmP94/CT was evaluated for diagnostic use, and it demonstrated high sensitivity and specificity when tested with the sera from mice experimentally infected with B. microti and closely related parasites. Moreover, the immunoprotective property of rBmP94/CT as a subunit vaccine was evaluated in BALB/c mice against a B. microti challenge, but no significant protection was observed. Our data suggest that the immunodominant antigen BmP94 could be a promising candidate for diagnostic use for human babesiosis.
通过免疫筛选 cDNA 表达文库，鉴定了一个编码微小巴贝斯虫 94kDa 蛋白的新基因 BmP94。全长的 BmP94 在大肠杆菌中表达（rBmP94），但以低产量形成不溶性形式，而基因的截断亲水 C 末端区域则作为可溶性蛋白（rBmP94/CT）表达，提高了产量。针对 rBmP94/CT 的抗血清通过 Western blot 分析识别寄生虫提取物中的 94kDa 天然蛋白。接下来，使用 rBmP94/CT 进行 ELISA 评估，用于诊断用途，用感染微小巴贝斯虫和密切相关寄生虫的实验感染小鼠的血清进行测试时，显示出高灵敏度和特异性。此外，rBmP94/CT 作为亚单位疫苗在 BALB/c 小鼠中针对微小巴贝斯虫的挑战进行了免疫保护特性评估，但未观察到显著保护。我们的数据表明，免疫显性抗原 BmP94 可能是人类巴贝斯虫病诊断用途的有前途的候选物。
