聚合酶链反应微孔板杂交法检测解脲脲原体及药敏分析

目的 建立敏感、特异的聚合酶链反应－微孔板杂交法（ＰＣＲ－ＭＰＨ）检测解脲脲原体（Ｕｕ）的感染，并分析药敏情况。方法 将带有生物素标记的Ｕｕ的聚合酶链反应产物结合在链霉亲和素包被的微孔板上，与标记地高辛的探针进行杂交后，通过标记碱性磷酸酶的抗地高辛抗体（ａｎｔｉ－ＤＩＧ－ＡＰ）与杂交分子进行反应，最终经底物显色后读数；同时应用生物梅里埃公司支原体培养试剂盒（ＩＳＴ）进行了Ｕｕ的培养及药敏试验。结果 通过优化多种试验条件建立了ＰＣＲ－ＭＰＨ法检测Ｕｕ，并分析了不同人群１５８份标本，其中微孔板杂交法阳性６５份，培养法阳性５６份；药敏结果显示，敏感率原始霉素１００％、交沙霉素９６．６％、强力霉素８９．７％、四环素７９．３％、氧氟沙星３４．５％、红霉素６．９％。结论 ＰＣＲ－ＭＰＨ法采用非放射性标记，具有无污染、经济、

Detectionof ureaplasma urealyticum by polymerase chain reaction-microplate hybridization andanti-Uu susceptibility test

Objectives To establish a sensitive and special method for the detection of Ureaplasma urealyticum(Uu) using PCR microplate hybridization (PCR MPH). Methods A primer of ureasea gene was labeled by biotin. The amplification product was captured on streptavidin coated microplates, then products were quantified by hybridization with a digoxigenin labeled internal oligonucleotide probe. After revelation with an anti digoxigenin alkaline phosphatase coupled antibody(anti DIG AP), the amount was determined by optical reading. At the same time, PCR MPH was compared with Bio Merieux Mycosplasma IST. Results A method of PCR MPH for detecting Uu DNA was established. The morbidity among three groups for detecting 158 clinical samples was analysed. 65 were detected by PCR MPH and 56 by culture.Conclusion The results showed that this assay is rapid, sensitive, specific, and accurate, and is of value in clinical therapy.