Impact of various handling and storage conditions on quantitative detection of hepatitis C virus RNA

doi:10.1016/S0168-8278(96)80116-4

Journal of Hepatology

Volume 25, Issue 3, September 1996, Pages 307-311

https://doi.org/10.1016/S0168-8278(96)80116-4 Get rights and content

Abstract

Background/Aims: Both HCV RNA viral load and HCV genotype have been described as important predicting factors determining the response to interferon in chronic hepatitis C. To investigate whether processing and storage conditions might influence the stability and could alter the concentration of the HCV RNA in serum, quantification of HCV RNA was performed by branched DNA assay.

Methods: We studied serum samples obtained from seven patients with histologically proven chronic hepatitis C. These were subjected to the following physical conditions: (1) immediate quantification, (2) storage at room temperature for 5 days, (3) storage at 4°C for 5 days, (4) storage at −20°C for 5 days, (5) storage at −80°C for 5 days, (6) five freeze-thaw cycles, (7) blood unspun for 4 h at room temperature then centrifuged and stored at −80°C for 5 days, (8) storage at 4°C for 6 months, (9) storage at −20°C for 6 months, (10) storage at −80°C for 6 months.

Results: A loss of 100% HCV RNA titers was observed after storage at RT for 5 days and then storage at 4°C for 6 months. A surprising decrease of HCV RNA titer (15.6%) was observed in sera stored for 5 days at −20°C. Five freeze-thaw cycles resulted in a 16% decrease of the HCV RNA level. When centrifugation was performed after a 4 h delay at room temperature, a significant loss of HCV RNA titers of 29.5% was observed. Long-term stability (6 months) was observed at −80°C with a slight loss of about of 10% HCV RNA titers, but a significant decrease in HCV RNA of 23% was observed at −20°C. The reproducibility of the bDNA assay on five patient samples was performed eight times in duplicate and showed an average coefficient of variation of 9.1%.

Conclusions: These data confirm the importance of storage and handling in measuring the amount of HCV RNA in clinical samples.

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Impact of various handling and storage conditions on quantitative detection of hepatitis C virus RNA

Abstract

Lancet

Lancet

Lancet

Anal Biochem

Lancet

Hepatology

Gastroenterology

Lancet

Isolation of a cDNA clone derived from a blood-borne non-A, non-B viral hepatitis genome

Science

An assay for circulating antibodies to a major etiologic virus of human non-A, non-B hepatitis

Science

Detection of hepatitis C virus RNA by a two-stage polymerase chain reaction with two pairs of primers deduced from the 5′-noncoding region

Jpn J Exp Med

Quantitation of hepatitis C virus RNA by competitive polymerase chain reaction

J Med Virol

Quantitative analysis of hepatitis C virus by multicyclic RT-PCR

Jpn J Gastroenterol

A novel method for the detection of hepatitis B virus in human serum samples without blotting or radioactivity

Branched DNA amplification multimers for the sensitive, direct detection of human hepatitis virus

Nucleic Acids Symp Series