CAJ | 학술논문

为探索一种快速提取血吸虫虫卵并保持虫卵抗原性的方法，本研究用血吸虫尾蚴感染兔并收集其肝脏，分别用不同浓度的NaOH、不同温度和时间消蚀含有虫卵的肝脏匀浆液，以确定最佳提取虫卵条件。通过方阵法确定所提取虫卵的可溶性抗原最佳包被浓度，并对虫卵抗原灵敏度进行了检测。另外，通过测定75份阳性血吸虫水牛血清和60份阴性水牛血清，进一步检测了此法提取并制备抗原的灵敏性。结果表明，本研究所建立的方法在不同NaOH浓度、作用温度和作用时间下，对提取的虫卵均产生影响。NaOH浓度越高，作用时间越久，消蚀温度越高，对虫卵的破坏越显著，表现在虫卵内容物的凝聚和消融，虫卵的空壳化增多。3种不同浓度NaOH 提取虫卵的抗原检测下限分别为1/800、1/1600、1/3200，较过筛法的抗原检测下限1/400有显著提高。对水牛的血清检测结果表明，用NaOH消蚀法提取的虫卵抗原检测血吸虫灵敏性是93%，比常规过筛法高5%，特异性是100%，比常规过筛法高3%。本研究结果表明NaOH消蚀法可快速提取高洁净度的虫卵并保持虫卵的抗原性不被破坏，该法提取虫卵的最佳NaOH浓度是5%~10%，作用温度是45℃左右，作用时间是5~10 min。在最佳条件下制备的虫卵较常规过筛法的纯净度高，虫卵得率也是常规过筛法的3倍以上。
위탐색일충쾌속제취혈흡충충란병보지충란항원성적방법，본연구용혈흡충미유감염토병수집기간장，분별용불동농도적NaOH、불동온도화시간소식함유충란적간장균장액，이학정최가제취충란조건。통과방진법학정소제취충란적가용성항원최가포피농도，병대충란항원령민도진행료검측。령외，통과측정75빈양성혈흡충수우혈청화60빈음성수우혈청，진일보검측료차법제취병제비항원적령민성。결과표명，본연구소건립적방법재불동NaOH농도、작용온도화작용시간하，대제취적충란균산생영향。NaOH농도월고，작용시간월구，소식온도월고，대충란적파배월현저，표현재충란내용물적응취화소융，충란적공각화증다。3충불동농도NaOH 제취충란적항원검측하한분별위1/800、1/1600、1/3200，교과사법적항원검측하한1/400유현저제고。대수우적혈청검측결과표명，용NaOH소식법제취적충란항원검측혈흡충령민성시93%，비상규과사법고5%，특이성시100%，비상규과사법고3%。본연구결과표명NaOH소식법가쾌속제취고길정도적충란병보지충란적항원성불피파배，해법제취충란적최가NaOH농도시5%~10%，작용온도시45℃좌우，작용시간시5~10 min。재최가조건하제비적충란교상규과사법적순정도고，충란득솔야시상규과사법적3배이상。
A simple and rapid method for isolating schistosome eggs was developed in the present study. Rabbits were infected with cercariaes and their livers were collected afterwards. The livers were homogenized and treated with NaOH at different concentrations, temperatures and times. The optimal concentration of soluble egg antigen (SEA) for ELISA use was determined by reacting with positive sera of schistosomiasis. Total 75 positive and 60 negative buffalo serum samples were tested to determine the reactivity of SEA extracts. Treatment of liver homogenates with high concentration of NaOH for a long time at the high temperature significantly reduced the reactivity of SEA by causing aggression and ablation of egg contents and increased rate of empty egg shells. The SEA products obtained from liver homogenates treated with 5%, 10%and 20%NaOH reacted in ELISA with positive serum samples diluted at 1:800, 1:1600 and 1:3200, which were lower than the SEA extracted from eggs by mesh method and diluted at 1:400. The results of ELISA showed that the sensitivity and specificity were 93%and 100%, which were 5%and 3%higher than the SEA obtained by mesh method. The optimal conditions to isolate eggs from livers were 5%-10%NaOH at 45℃for 5-10min. The yield of eggs was almost three times more than conventional mesh method. These results indicated that a large amount of pure eggs could be isolated without loss of antigenicity by using simple and rapid NaOH-maceration method.