Highly reproducible bactericidal activity test results by using a modified national committee for clinical laboratory standards broth macrodilution technique
Highly reproducible bactericidal activity test results by using a modified national committee for clinical laboratory standards broth macrodilution technique
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DOI：
10.1128/jcm.37.6.1881-1884.1999
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发表时间：
1999-06-01
影响因子：
9.4
通讯作者：
Peterson, LR
中科院分区：
文献类型：
--
作者：
Hacek, DM;Dressel, DC;Peterson, LR
历史上，即使采用先前的标准化方法，杀菌测试也表现出可变的再现性。建议对美国临床实验室标准化委员会（NCCLS）肉汤大量稀释法进行几项修改，以提高重现性，建议对已批准的NCCLS指南进行修改（M21-A和M26-A）包括省略Mueller-Hinton肉汤的血清补充，在35 ℃下孵育管24小时，不搅拌直至取样，用6个稀释液而不是9个稀释液重复进行所有试验，再孵育试验24 h以解决不一致的杀菌活性试验结果，使用每个透明试管中的单个0.1 ml样品进行传代培养，并采用替代方法计算终点测定。为了在临床实验室环境中测试这些建议，我们在224项单独的杀菌活性测试中使用了改良的方法。进行了102次血清杀菌效价（SBT）和122次最低杀菌浓度（MBC）测定。通过将重现性定义为重复检测+/- 1稀释度之间的一致性，我们发现224次检测中有207次（92%）在24小时传代培养点具有重现性（SET试验为94%，MBC试验为91%），当将具有不一致结果的17项试验再孵育24小时进行第二次传代培养时，224次检测中只有1次（0.4%）仍存在差异。所使用的方法对于选择进行杀菌活性测试的临床实验室是实用的，并确保重复测定之间的高水平再现性。一次测试的总费用约为25美元。
Bactericidal testing historically has exhibited variable reproducibility, even when prior standardized methods were employed. Several modifications to the National Committee for Clinical Laboratory Standards (NCCLS) broth macrodilution method are proposed to improve reproducibility, Recommended changes from the approved NCCLS guidelines (M21-A and M26-A) include omitting serum supplementation of Mueller-Hinton broth, incubating tubes at 35 degrees C for 24 h with no agitation until they are sampled, running all tests in duplicate with six dilutions instead of nine, reincubating the test for an additional 24 h to resolve discrepant bactericidal activity test results, using a single 0.1-ml sample from each clear tube for subculture, and adopting an alternate method for calculating endpoint determination. In order to test these recommendations in a clinical laboratory setting, we used the modified methodology on 224 separate tests for bactericidal activity. There were 102 serum bactericidal titer (SBT) and 122 minimum bactericidal concentration (MBC) assays performed. By defining reproducibility as agreement between duplicate tests +/- 1 dilution, we found 207 of 224 tests (92%) were reproducible at the 24-h subculture point (94% for the SET assay and 91% for the MBC assay), When the 17 assays with discrepant results were incubated an additional 24 h for a second subculture, only 1 of 224 tests (0.4%) remained discrepant. The method used is practical for a clinical laboratory that chooses to perform bactericidal activity testing and assures a high level of reproducibility between duplicate assays. The total cost of a test was approximately $25.00.