Inhibitory effects of urine on the polymerase chain reaction for cytomegalovirus DNA.
Author(s): Khan G, Kangro HO, Coates PJ, Heath RB
Publication: J Clin Pathol, 1991, Vol. 44, Page 360-5
PubMed ID: 1646235 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
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Study Purpose
The purpose of this study was to identify the urine component that inhibited PCR detection of CMV.
Summary of Findings:
The authors report that urea is a complete inhibitor of PCR at a concentration of 100 mM and partially inhibitory at a concentration of 50 mM. The inhibitory activity of urine was independent of creatinine and electrolyte concentrations. Two neonatal specimens with extremely low urea concentrations (10 mM) that were reactive for glucose also resulted in inhibition of the PCR reaction.
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Not specified
Platform:
Analyte Technology Platform DNA PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Aliquots and Components Biospecimen components Urea concentration (5-250 mM)
Creatinine concentration
Na+ concentration
K+ concentration
Glucose concentration
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Study Purpose
The purpose of this study was to identify purification and analytical methods that circumvent the inhibitory effect of urine for PCR analysis.
Summary of Findings:
While the authors report that both dialysis and ultrafiltration of urine specimens were successful in the removal of inhibitory activity, Millipore cups were preferred due to benefits associated with volume, time, and financial requirements. Specimen filtration, however, reduced amplicon intensity, which was circumvented through the use of a nested two stage PCR approach that increased sensitivity 10-fold compared to conventional one stage PCR.
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Not specified
Platform:
Analyte Technology Platform DNA PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Analyte Extraction and Purification Analyte purification Dialysis
Centricon-100 ultrafiltration tubes
Millipore ultrafiltration cups
PCR Specific Technology platform One stage PCR
Two stage nested PCR