Feng Shaw-Huey, Tsai Shien, Rodriguez Jose, Newsome Tamara, Emanuel Peter, Lo Shyh-Ching
Department of Environmental and Infectious Disease Sciences, Armed Forces Institute of Pathology (AFIP), Washington, DC, USA.
Hybridoma (Larchmt). 2006 Aug;25(4):193-201. doi: 10.1089/hyb.2006.25.193.
Burkholderia mallei and B. pseudomallei are designated category B biothreat agents on the "select agents" list established by the NIH and CDC. Development of monoclonal antibodies (MAbs) that could effectively differentiate these two closely related species of bacteria and other non-pathogenic Burkholderia bacteria is urgently needed. Splenocytes from mice immunized with various antigen preparations from either B. mallei (American Type Culture Collection [ATCC] 23344) or B. pseudomallei (ATCC 23343) were used for production of hybridomas. Using a three-step cross-screening protocol, a total of 10 hybridomas were selected that produced MAbs which specifically recognized B. mallei 23344 but did not bind B. pseudomallei, Pseudomonas aeruginasa, or any of the other nine Burkholderia species tested. All 10 MAbs targeted to the lipopolysaccharide (LPS) molecules of B. mallei and reacted strongly with 12 out of 15 different strains of B. mallei tested. A total of 14 hybridomas that produced MAbs reacting with B. pseudomallei 23343, but not with B. mallei, P. aeruginasa, or any other nine non-pathogenic Burkholderia species were also selected. All 14 MAbs appeared to react with a proteinase K-sensitive 200-kDa band by immunoblotting analysis. Surprisingly, these 14 MAbs that were raised against the ATCC 23343 strain failed to react to any of the other 13 different strains of B. pseudomallei examined. In conclusion, our B. mallei-specific MAbs can effectively recognize 80% of the different B. mallei strains tested, and all the B. pseudomallei-specific MAbs appeared to react with a unique antigen present only in the ATCC 23343 strain, but not in any other strains of B. pseudomallei tested.
鼻疽伯克霍尔德菌和类鼻疽伯克霍尔德菌被美国国立卫生研究院（NIH）和疾病控制与预防中心（CDC）列入“特定病原体”名单中的B类生物威胁因子。迫切需要开发能够有效区分这两种密切相关的细菌物种以及其他非致病性伯克霍尔德菌的单克隆抗体（MAb）。用来自鼻疽伯克霍尔德菌（美国模式培养物集存库[ATCC] 23344）或类鼻疽伯克霍尔德菌（ATCC 23343）的各种抗原制剂免疫的小鼠脾细胞用于制备杂交瘤。使用三步交叉筛选方案，共选择了10个杂交瘤，它们产生的单克隆抗体能特异性识别鼻疽伯克霍尔德菌23344，但不与类鼻疽伯克霍尔德菌、铜绿假单胞菌或所测试的其他9种伯克霍尔德菌物种结合。所有10种单克隆抗体均靶向鼻疽伯克霍尔德菌的脂多糖（LPS）分子，并与所测试的15种不同鼻疽伯克霍尔德菌菌株中的12种强烈反应。还选择了总共14个产生与类鼻疽伯克霍尔德菌23343反应但不与鼻疽伯克霍尔德菌、铜绿假单胞菌或其他9种非致病性伯克霍尔德菌物种反应的单克隆抗体的杂交瘤。通过免疫印迹分析，所有14种单克隆抗体似乎都与一条对蛋白酶K敏感的200 kDa条带反应。令人惊讶的是，这些针对ATCC 23343菌株产生的14种单克隆抗体未能与所检测的其他13种不同类鼻疽伯克霍尔德菌菌株中的任何一种反应。总之，我们的鼻疽伯克霍尔德菌特异性单克隆抗体可以有效识别80%所测试的不同鼻疽伯克霍尔德菌菌株，并且所有类鼻疽伯克霍尔德菌特异性单克隆抗体似乎都与仅存在于ATCC 23343菌株中而非所测试的任何其他类鼻疽伯克霍尔德菌菌株中的一种独特抗原反应。
