中华检验医学杂志
中華檢驗醫學雜誌
중화검험의학잡지
CHINESE JOURNAL OF LABORATORY MEDICINE
2010年
9期
884-890
,共7页
江凌晓%王艳芳%郝卫%丘立文%蔡建飘%潘玉先%陈文霞%姜长宏%林丽娟%车小燕
江凌曉%王豔芳%郝衛%丘立文%蔡建飄%潘玉先%陳文霞%薑長宏%林麗娟%車小燕
강릉효%왕염방%학위%구립문%채건표%반옥선%진문하%강장굉%림려연%차소연
曲霉%感染%诊断%标志物
麯黴%感染%診斷%標誌物
곡매%감염%진단%표지물
Aspergillus%Iinfection%Diagnosis%Biomarker
目的 筛选出可用于侵袭性曲霉感染实验室早期诊断的特异性单克隆抗体.方法 应用单克隆抗体技术制备针对不同曲霉抗原的特异性抗曲霉单克隆抗体,根据单抗识别表位不同,初步筛选出两组灵敏度高、特异性强的单抗组合,并分别建立了双抗体夹心ELISA检测法.通过检测常见临床和环境分离曲霉株、马尔尼菲青霉及念珠菌培养液,感染动物模型标本,临床标本,初步评估检测方法的敏感性和特异性.并应用WB对抗体所识别靶标进行初步鉴定.结果 获得32株稳定分泌曲霉单抗的杂交瘤细胞株,建立了2种双抗体夹心ELISA法,第1种方法可识别环境和临床分离的19种曲霉.第2种方法仅特异性识别临床和环境分离的烟曲霉,与其他曲霉抗原无交叉.对同一种烟曲霉培养液而言,第1种方法可检测稀释度为第2种方法的10倍.WB分析显示该组单抗识别的靶标为相对分子质量在25 000～75 000之间的甘露聚糖蛋白.结论 本研究获得了可用于侵袭性曲霉感染实验室诊断的特异性单克隆抗体,单抗识别的抗原为相对分子质量在25 000～75 000之间的甘露聚糖蛋白.该抗原是侵袭性曲霉感染早期诊断的潜在标志物.
目的 篩選齣可用于侵襲性麯黴感染實驗室早期診斷的特異性單剋隆抗體.方法 應用單剋隆抗體技術製備針對不同麯黴抗原的特異性抗麯黴單剋隆抗體,根據單抗識彆錶位不同,初步篩選齣兩組靈敏度高、特異性彊的單抗組閤,併分彆建立瞭雙抗體夾心ELISA檢測法.通過檢測常見臨床和環境分離麯黴株、馬爾尼菲青黴及唸珠菌培養液,感染動物模型標本,臨床標本,初步評估檢測方法的敏感性和特異性.併應用WB對抗體所識彆靶標進行初步鑒定.結果 穫得32株穩定分泌麯黴單抗的雜交瘤細胞株,建立瞭2種雙抗體夾心ELISA法,第1種方法可識彆環境和臨床分離的19種麯黴.第2種方法僅特異性識彆臨床和環境分離的煙麯黴,與其他麯黴抗原無交扠.對同一種煙麯黴培養液而言,第1種方法可檢測稀釋度為第2種方法的10倍.WB分析顯示該組單抗識彆的靶標為相對分子質量在25 000～75 000之間的甘露聚糖蛋白.結論 本研究穫得瞭可用于侵襲性麯黴感染實驗室診斷的特異性單剋隆抗體,單抗識彆的抗原為相對分子質量在25 000～75 000之間的甘露聚糖蛋白.該抗原是侵襲性麯黴感染早期診斷的潛在標誌物.
목적 사선출가용우침습성곡매감염실험실조기진단적특이성단극륭항체.방법 응용단극륭항체기술제비침대불동곡매항원적특이성항곡매단극륭항체,근거단항식별표위불동,초보사선출량조령민도고、특이성강적단항조합,병분별건립료쌍항체협심ELISA검측법.통과검측상견림상화배경분리곡매주、마이니비청매급념주균배양액,감염동물모형표본,림상표본,초보평고검측방법적민감성화특이성.병응용WB대항체소식별파표진행초보감정.결과 획득32주은정분비곡매단항적잡교류세포주,건립료2충쌍항체협심ELISA법,제1충방법가식별배경화림상분리적19충곡매.제2충방법부특이성식별림상화배경분리적연곡매,여기타곡매항원무교차.대동일충연곡매배양액이언,제1충방법가검측희석도위제2충방법적10배.WB분석현시해조단항식별적파표위상대분자질량재25 000～75 000지간적감로취당단백.결론 본연구획득료가용우침습성곡매감염실험실진단적특이성단극륭항체,단항식별적항원위상대분자질량재25 000～75 000지간적감로취당단백.해항원시침습성곡매감염조기진단적잠재표지물.
Objective To screen monoclonal antibodies (mAbs) for early diagnosis of invisive Aspergillus. Methods Monoclonal antibodies against different antigens of Aspergillus fumigatus were produced. The two pairs of combinations of monoclonal antibodies were selected accoring the distinct epitopes and double-antibody sandwich ELISA based on mAbs above were established. The sensitivity and specificity of the methods were analyzed by detecting culture supernatants of clinical isolates and environmental isolatesof Aspergillus. spp, Penicillium Marneffei, Candidas, and serum from animal models and patients. The epitopes recognized by mAbs were identified by immunobotting. Results A total of 32 hybridoma cell lines that stably produced MAbs were obtained. Two double- antibody sandwich ELISAs were established. One method was specific for 19 clinical isolates and environmental isolates of Aspergillus. spp, whereas the other one was specific for the clinical and environmental isolates of Aspergillus fumigatus without cross-reation with other Aspergillus. spp. For the same kind of medium of Aspergillus fumigatus, the sensitivity of the first method was 10 fold higher than the second method. Conclusions The specific mAbs for early diagnosis of invisive Aspergillus were obtained. Antigen recognized by the specific mAbs was mannoprotein with molecular weights of approximately 25 000-75 000. This antigen was potential early diagnostic marker for invasive Aspergillus.
