| 摘要: |
| 目的 建立简便有效的样本处理方法，提高实时荧光定量PCR法检测尿液中BK病毒载量的敏感性和准确性。方法 任意选择24例BK病毒阳性样本，每一样本进行4种不同的处理：原尿、原尿病毒DNA提纯、110稀释尿液、1100稀释尿液，实时荧光定量PCR法检测各组样本BK病毒载量，所测数据用SPSS 11.0进行统计学分析。结果 4种不同的处理方法对实时荧光定量PCR法检测尿液中BK病毒载量有明显的影响：原尿阴性3例，病毒载量在4组中的最低值占66.7%；1100稀释尿阴性2例，病毒载量在4组中的最高值占79.2%，但中位值和110稀释组差异无统计学意义；DNA纯化组与110稀释尿液病毒检出率相当，均为100%，但DNA纯化组目标基因的流失量较明显。结论 在临床尿液中BK病毒载量检测时，110稀释尿在实时荧光定量PCR检测BK病毒时DNA损失小，效率高，且处理过程简便、成本低，是一种较好的尿样本处理方法。 |
| 关键词: BK病毒 尿 实时荧光定量PCR 病毒载量 |
| DOI：10.3724/SP.J.1008.2010.0438 |
| 投稿时间：2009-11-27修订日期：2010-03-12 |
| 基金项目:广东省自然科学基金自由项目（04010491）. |
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| Optimization of urinary sample preparation process for real-time PCR detection of BK virus |
| LIN Chun-lan1, LU Yu-hong2*, CHEN Shao-hua3, LI Yang-qiu3, ZHU Kang-er2, ZHANG Tao2 |
| (1. Department of Biochemistry, Medical College of Jinan University, Guangzhou 510632, Guangdong, China;2. Hematological Station, the First Affiliated Hospital of Jinan University, Guangzhou 510632, Guangdong, China;3. Hematology Institute of Medical College, Jinan University, Guangzhou 510632, Guangdong, China) |
| Abstract: |
| Objective To develop an effective preparation method to improve the sensitivity and accuracy of real-time PCR in detection of BK virus’s (BKV’s) load in urine samples. Methods A total of 24 samples documented as positive probes in primary detection were enrolled in this study. The candidate samples were prepared by 4 different approaches: unprocessed urine, BKV’s DNA extracted from urine, 110 diluted urine, and 1100 diluted urine; and then they were subjected to real-time PCR examination to obtain the viral load. The data obtained were analyzed by SPSS 11.0. Results The four different preparation processes for urinary specimens had significant impact on detection results of real-time PCR. Three samples were negative in the unprocessed urine group and 66.7% of its samples had the lowest viral loads compared with the other three groups. Two samples in the 1100 diluted urine group were negative and 79.2% of its samples had the highest viral loads, but its median load was similar to that of the 110 group. Viral gene was detected in all samples in the DNA extraction group and 110 diluted urine group, but the loss of the target gene was more severe in the DNA extraction group. Conclusion The 110 diluted urine is better for real-time PCR detection of BKV’s load, as it lose less viral gene and is more efficient, easy to perform and economical. |
| Key words: BK virus urine real-time PCR viral load |
