Han Eun-Taek, Watanabe Risa, Sattabongkot Jetsumon, Khuntirat Benjawan, Sirichaisinthop Jeeraphat, Iriko Hideyuki, Jin Ling, Takeo Satoru, Tsuboi Takafumi
Cell-Free Science and Technology Research Center, Ehime University, Matsuyama, Ehime 790-8577, Japan.
J Clin Microbiol. 2007 Aug;45(8):2521-8. doi: 10.1128/JCM.02117-06. Epub 2007 Jun 13.
Loop-mediated isothermal amplification (LAMP), a novel nucleic acid amplification method, was developed for the clinical detection of four species of human malaria parasites: Plasmodium falciparum, P. vivax, P. malariae, and P. ovale. We evaluated the sensitivity and specificity of LAMP in comparison with the results of microscopic examination and nested PCR. LAMP showed a detection limit (analytical sensitivity) of 10 copies of the target 18S rRNA genes for P. malariae and P. ovale and 100 copies for the genus Plasmodium, P. falciparum, and P. vivax. LAMP detected malaria parasites in 67 of 68 microscopically positive blood samples (sensitivity, 98.5%) and 3 of 53 microscopically negative samples (specificity, 94.3%), in good agreement with the results of nested PCR. The LAMP reactions yielded results within about 26 min, on average, for detection of the genus Plasmodium, 32 min for P. falciparum, 31 min for P. vivax, 35 min for P. malariae, and 36 min for P. ovale. Accordingly, in comparison to the results obtained by microscopy, LAMP had a similar sensitivity and a greater specificity and LAMP yielded results similar to those of nested PCR in a shorter turnaround time. Because it can be performed with a simple technology, i.e., with heat-treated blood as the template, reaction in a water bath, and inspection of the results by the naked eye because of the use of a fluorescent dye, LAMP may provide a simple and reliable test for routine screening for malaria parasites in both clinical laboratories and malaria clinics in areas where malaria is endemic.
环介导等温扩增技术（LAMP）是一种新型核酸扩增方法，用于临床检测四种人类疟原虫：恶性疟原虫、间日疟原虫、三日疟原虫和卵形疟原虫。我们将LAMP的灵敏度和特异性与显微镜检查和巢式PCR的结果进行了比较评估。LAMP对三日疟原虫和卵形疟原虫的靶标18S rRNA基因的检测限（分析灵敏度）为10拷贝，对疟原虫属、恶性疟原虫和间日疟原虫为100拷贝。LAMP在68份显微镜检查呈阳性的血样中的67份中检测到疟原虫（灵敏度为98.5%），在53份显微镜检查呈阴性的血样中的3份中检测到疟原虫（特异性为94.3%），与巢式PCR的结果高度一致。LAMP反应平均约26分钟即可得出疟原虫属的检测结果，恶性疟原虫为32分钟，间日疟原虫为31分钟，三日疟原虫为35分钟，卵形疟原虫为36分钟。因此，与显微镜检查结果相比，LAMP具有相似的灵敏度和更高的特异性，并且在更短的周转时间内得出与巢式PCR相似的结果。由于LAMP可以采用简单的技术进行，即以热处理后的血液为模板、在水浴中进行反应以及由于使用荧光染料而可通过肉眼检查结果，因此LAMP可为疟疾流行地区的临床实验室和疟疾诊所进行疟原虫的常规筛查提供一种简单可靠的检测方法。
