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目的：建立操作简便、结果客观的微孔板杂交－ 酶联显色检测土拉弗朗西斯氏菌 Ｐ Ｃ Ｒ 产物的技术。方法：将 Ｐ Ｃ Ｒ 产物或产物的克隆作为捕获探针包被聚乙烯微孔板， Ｐ Ｃ Ｒ 引物５′末端生物素标记，扩增后用作探针杂交，通过比较影响微孔板杂交的包被、杂交和显色等因素，建立 Ｐ Ｃ Ｒ－ Ｅ Ｉ Ａ 技术。结果：通过比较包被缓冲液发现镁离子及其离子强度是影响 Ｄ Ｎ Ａ 包被的重要因素；包被的 Ｄ Ｎ Ａ 以线型质粒较佳，每孔包被３００ ｎｇ 左右的 Ｄ Ｎ Ａ 杂交效果最好；杂交液中加和不加甲酰胺对杂交影响不大；用链霉亲和素化的辣根过氧化物酶显色均能达到检测杂交体的目的，结论：我们确定了 Ｐ Ｃ Ｒ－ Ｅ Ｉ Ａ 技术的最佳包被、杂交和显色的条件，该方法的建立为 Ｐ Ｃ Ｒ 真正用于临床实验奠定了基础。
OBJECTIVE: To establish a simple and objective method for the determination of P C R product of Francisella tularensis by microplate hybridization and enzyme-linked chromotagraphy. Methods: The clones of PCR products or products were used as capture probe to coat polyethylene microplate. The PCR primers were labeled with biotin at the 5 ’end and amplified for probe hybridization. Microplate hybridization Coating, hybridization and color development and other factors, the establishment of P C R-E I A technology. Results: The magnesium ions and their ionic strength were found to be important factors affecting the D N A coating by comparing the coating buffer. The coated D N A with linear plasmids was preferably coated with about 300 ng D N A per well Hybridization effect is best; hybridization liquid with and without formamide on hybridization has little effect; with streptavidin horseradish peroxidase color detection hybrids can achieve the purpose, we conclude: we have identified The optimal coating, hybridization, and chromogenic conditions for P C R-E I A technology have laid the foundation for P C R’s true use in clinical trials.