ABSTRACT
Objective To optimize the application of the nested PCR method for the detection of malaria according to the working practice,so as to improve the efficiency of malaria detection. Methods Premixing solution of PCR,internal primers for further amplification and new designed primers that aimed at two Plasmodium ovale subspecies were employed to optimize the reaction system,reaction condition and specific primers of P. ovale on basis of routine nested PCR. Then the specificity and the sensitivity of the optimized method were analyzed. The positive blood samples and examination samples of malaria were detected by the routine nested PCR and the optimized method simultaneously,and the detection results were compared and analyzed. Re-sults The optimized method showed good specificity,and its sensitivity could reach the pg to fg level. The two methods were used to detect the same positive malarial blood samples simultaneously,the results indicated that the PCR products of the two methods had no significant difference,but the non-specific amplification reduced obviously and the detection rates of P. ovale subspecies improved,as well as the total specificity also increased through the use of the optimized method. The actual detection results of 111 cases of malarial blood samples showed that the sensitivity and specificity of the routine nested PCR were 94.57％and 86.96％,respectively,and those of the optimized method were both 93.48％,and there was no statistically significant dif-ference between the two methods in the sensitivity(P>0.05),but there was a statistically significant difference between the two methods in the specificity(P<0.05). Conclusion The optimized PCR can improve the specificity without reducing the sensi-tivity on the basis of the routine nested PCR,it also can save the cost and increase the efficiency of malaria detection as less ex-periment links.