INHIBITORY EFFECTS OF URINE ON THE POLYMERASE CHAIN-REACTION FOR CYTOMEGALOVIRUS DNA
INHIBITORY EFFECTS OF URINE ON THE POLYMERASE CHAIN-REACTION FOR CYTOMEGALOVIRUS DNA
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DOI：
10.1136/jcp.44.5.360
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发表时间：
1991-05-01
影响因子：
3.4
通讯作者：
HEATH, RB
中科院分区：
文献类型：
--
作者：
KHAN, G;KANGRO, HO;HEATH, RB
研究了取自新生儿和年龄较大儿童的尿液样本（其中一些已知感染了巨细胞病毒）对聚合酶链反应（PCR）的抑制作用。尿素是尿液中的主要抑制成分，浓度超过 50 mM 时会抑制 PCR。大龄儿童的尿液样本比新生儿的尿液样本更具抑制性。这与大龄儿童尿样中尿素浓度高于新生儿尿样相关。然而，尽管尿素浓度较低，但 13 份新生儿尿液样本中的 2 份仍具有抑制作用，这可能是由于肠外营养产生的代谢产物所致。通过简单的透析或超滤即可有效消除尿液的抑制作用。通过使用“嵌套”引物和修改后的 PCR 方案（需要在两阶段 50 个循环 PCR 的前 20 个循环中使用减少的反应物），进一步提高了 PCR 检测尿液中巨细胞病毒 DNA 的灵敏度和特异性。
The inhibitory effects of urine samples taken from neonates and older children, some of which were known to be infected with cytomegalovirus, on the polymerase chain reaction (PCR) were investigated. Urea was the major inhibitory component of urine and inhibited the PCR at a concentration of more than 50 mM. Urine samples from older children were more inhibitory than those from neonates. This correlated with the higher concentration of urea generally found in urine samples from older children compared with neonatal urines. Two of 13 neonatal urine samples, however, were inhibitory despite low urea concentrations-presumably due to metabolites derived from parenteral nutrition. The inhibitory effects of urine were effectively removed by simple dialysis or ultrafiltration. The sensitivity and specificity of PCR for detecting cytomegalovirus DNA in urine were further improved by using "nested" primers and a modified PCR protocol entailing the use of reduced reactants in the first 20 cycles of a two-stage 50 cycle PCR.