Quantification of HIV-1 RNA load by one-tube-one-step RT PCR and real time PCR assay with TaqMan probe.

Watanaveeradej, Veerachai; Sirirattanaphoomee, Sudalak; Chantratita, Wasun; Nitayaphan, Sorachai; Viputtikul, Kwanjai; Kerdpanich, Angkool; Samakoses, Rudiwilai; Simasathien, Sriluk

Quantification of HIV-1 RNA load by one-tube-one-step RT PCR and real time PCR assay with TaqMan probe.

Date

2005-11-25

Authors

Watanaveeradej, Veerachai

Sirirattanaphoomee, Sudalak

Abstract

OBJECTIVES: To develop a less expensive assay to calculate HIV-1 viral load for use in resource-limited countries. MATERIAL AND METHOD: An In-house One-tube-one-step Viral Load Assay (IOVA) was developed by using real-time PCR-based with TaqMan probe. Primers and probe were designed from the conserved region of sequences from all HIV subtypes. A standard curve was generated from reference virus in various dilutions. IOVA was applied on 105 HIV-positive and 25 HIV-negative samples and compared with the results from ROCHE AMPLICLOR. RESULTS: IOVA measured HIV RNA in the samples ranging from 125 to 2 x 10(6) copies/mL. The coefficient of variation of intra- and inter-assay ranged from 0.68% to 7.89%. The sensitivity, specificity, positive and negative predictive values were 92%, 100%, 100% and 79.5% respectively. The parallel quantitative analysis showed high correlation (r=0.95) between IOVA and AMPLICOR. CONCLUSION: A new HIV-1 viral load assay was developed and validated. It was reliable and less expensive.

Description

Chotmaihet Thangphaet.

Citation

Watanaveeradej V, Sirirattanaphoomee S, Chantratita W, Nitayaphan S, Viputtikul K, Kerdpanich A, Samakoses R, Simasathien S. Quantification of HIV-1 RNA load by one-tube-one-step RT PCR and real time PCR assay with TaqMan probe. Journal of the Medical Association of Thailand. 2005 Nov; 88 Suppl 3(): S206-13