BACKGROUND: Comparison and evaluation of molecular diagnostic assays for the detection and quantification of hepatitis C virus (HCV) RNA have been limited by the lack of RNA controls and calibrators. Armored RNA technology is a means for producing RNA that is completely protected from plasma ribonucleases. This method produces recombinant pseudoviral particles that are noninfectious and contain predefined RNA sequences. METHODS: A consensus 412-base sequence from the 5'NCR/Core region of HCV subtype 2b was derived from 34 individually sequenced HCV genotype 2b variants. A DNA fragment encoding the consensus HCV-2b sequence was synthesized de novo, cloned, and expressed as an Armored RNA control. The resulting HCV-2b Armored RNA (AR-HCV-2b) contained the complete HCV-2b consensus RNA sequence encapsidated within a protective protein coat. RESULTS: AR-HCV-2b was fully recoverable from human plasma incubated at 4 degrees C for >300 days. The particles were tested in three clinical assay formats: Amplicor HCV Monitor 1.0, Quantiplex HCV RNA 2.0, and INNO-LiPA HCV II. When added into seronegative, nonviremic plasma, AR-HCV-2b showed reproducible signals and linear dilutions in both the Amplicor and Quantiplex assays. AR-HCV-2b was correctly identified as subtype 2b in the INNO-LiPA line probe assay. CONCLUSION: The HCV-2b Armored RNA control is a versatile, durable, ribonuclease-resistant viral RNA control that is compatible in three different clinical assay formats.
BACKGROUND: Comparison and evaluation of molecular diagnostic assays for the detection and quantification of hepatitis C virus (HCV) RNA have been limited by the lack of RNA controls and calibrators. Armored RNA technology is a means for producing RNA that is completely protected from plasma ribonucleases. This method produces recombinant pseudoviral particles that are noninfectious and contain predefined RNA sequences. METHODS: A consensus 412-base sequence from the 5'NCR/Core region of HCV subtype 2b was derived from 34 individually sequenced HCV genotype 2b variants. A DNA fragment encoding the consensus HCV-2b sequence was synthesized de novo, cloned, and expressed as an Armored RNA control. The resulting HCV-2b Armored RNA (AR-HCV-2b) contained the complete HCV-2b consensus RNA sequence encapsidated within a protective protein coat. RESULTS: AR-HCV-2b was fully recoverable from human plasma incubated at 4 degrees C for >300 days. The particles were tested in three clinical assay formats: Amplicor HCV Monitor 1.0, Quantiplex HCV RNA 2.0, and INNO-LiPA HCV II. When added into seronegative, nonviremic plasma, AR-HCV-2b showed reproducible signals and linear dilutions in both the Amplicor and Quantiplex assays. AR-HCV-2b was correctly identified as subtype 2b in the INNO-LiPA line probe assay. CONCLUSION: The HCV-2b Armored RNA control is a versatile, durable, ribonuclease-resistant viral RNA control that is compatible in three different clinical assay formats.
Authors: J D Callahan; S J Wu; A Dion-Schultz; B E Mangold; L F Peruski; D M Watts; K R Porter; G R Murphy; W Suharyono; C C King; C G Hayes; J J Temenak Journal: J Clin Microbiol Date: 2001-11 Impact factor: 5.948
Authors: Nick A Antonishyn; Vivian M Ast; Ryan R McDonald; Rabindra K Chaudhary; Lisa Lin; Anton P Andonov; Greg B Horsman Journal: J Clin Microbiol Date: 2005-10 Impact factor: 5.948
Authors: Joseph C Phan; Barrett J Nehilla; Selvi Srinivasan; Robert W Coombs; Kim A Woodrow; James J Lai Journal: Pharm Res Date: 2016-07-11 Impact factor: 4.200
Authors: John B A Okello; Linda Rodriguez; Debi Poinar; Kirsten Bos; Andrew L Okwi; Gabriel S Bimenya; Nelson K Sewankambo; Kenneth R Henry; Melanie Kuch; Hendrik N Poinar Journal: PLoS One Date: 2010-11-10 Impact factor: 3.240
Authors: G Colucci; J Ferguson; C Harkleroad; S Lee; D Romo; S Soviero; J Thompson; M Velez; A Wang; Y Miyahara; S Young; C Sarrazin Journal: J Clin Microbiol Date: 2007-09-26 Impact factor: 5.948