A sample preparation method which facilitates detection of bacteria in blood cultures by the polymerase chain reaction
A sample preparation method which facilitates detection of bacteria in blood cultures by the polymerase chain reaction
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DOI：
10.1016/s0167-7012(98)00015-3
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发表时间：
1998-05-01
影响因子：
2.2
通讯作者：
Radstrom, Peter
中科院分区：
文献类型：
--
作者：
Al-Soud, Waleed Abu;Lantz, Par-Gunnar;Radstrom, Peter
开发了一种样品制备方法，以消除血液培养物中 PCR 抑制剂的影响，并在 PCR 之前浓缩细菌细胞。该方法基于稀释、离心、NaOH 洗涤和添加牛血清白蛋白的组合。通过将肺炎链球菌和脑膜炎奈瑟氏菌添加到含有全血的培养基中来评估该样品制备的效率。两种细菌的检测水平均为每毫升血液培养基1 CFU。如果没有这种处理，当将 10μl 含有人全血的培养基直接添加到 PCR 混合物中时，PCR 无法检测到每个反应管 107 CFU 的肺炎链球菌。当用每ml接种有约0.3CFU肺炎链球菌的全血培养基并在37℃下培养来评估样品制备时，8小时后检测到阳性PCR产物。
A sample preparation method was developed to remove the effect of PCR inhibitors in blood cultures, and to concentrate the bacterial cells prior to PCR. This method was based on a combination of dilution, centrifugation, washing with NaOH and the addition of bovine serum albumin. The efficiency of this sample preparation was evaluated with S. pneumoniae and N. meningitidis added to a culture medium containing whole blood. The detection level of both bacteria was 1 CFU per ml of blood culture medium. Without this treatment, the PCR could not detect 107 CFU of S. pneumoniae per reaction tube when 10 mul of culture medium containing human whole blood was added directly to the PCR mixture. When the sample preparation was evaluated with culture medium containing whole blood inoculated with approximately 0.3 CFU of S. pneumoniae per ml and incubated at 37degreeC, a positive PCR product was detected after 8 h.