Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation

A semiautomated extraction protocol of HCV-RNA using Favorgen RNA extraction kit has been developed. The kit provided protocol was modified by replacing manual spin steps with vacuum filtration. The assay performance was evaluated by real-time qPCR based on Taqman technology. Assay linearity was con...

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Autores principales: Amin, Imran, Jabbar, Tania, Niazi, Fawad, Akhtar, Muhammad Saeed
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Hindawi Publishing Corporation 2015
Materias:
Research Article
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4383304/
https://www.ncbi.nlm.nih.gov/pubmed/25873968
http://dx.doi.org/10.1155/2015/367801
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author Amin, Imran
Jabbar, Tania
Niazi, Fawad
Akhtar, Muhammad Saeed
author_facet Amin, Imran
Jabbar, Tania
Niazi, Fawad
Akhtar, Muhammad Saeed
author_sort Amin, Imran
collection PubMed
description A semiautomated extraction protocol of HCV-RNA using Favorgen RNA extraction kit has been developed. The kit provided protocol was modified by replacing manual spin steps with vacuum filtration. The assay performance was evaluated by real-time qPCR based on Taqman technology. Assay linearity was confirmed with the serial dilutions of RTA (Turkey) containing 1 × (10(6), 10(5), 10(4), and 10(3)) IU mL(−1). Comparison of test results obtained by two extraction methods showed a good correlation (r = 0.95, n = 30) with detection limit of 10(2) IU mL(−1). The semiautomated vacuum filtration based protocol demonstrated high throughput: 35 minutes for the extraction of a batch of 30 samples (150 µL each) with reduced labor, time, waste, and cost. Performance characteristics of semiautomated system make it suitable for use in diagnostic purpose and viral load determinations.
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spelling pubmed-43833042015-04-13 Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation Amin, Imran Jabbar, Tania Niazi, Fawad Akhtar, Muhammad Saeed Int J Anal Chem Research Article A semiautomated extraction protocol of HCV-RNA using Favorgen RNA extraction kit has been developed. The kit provided protocol was modified by replacing manual spin steps with vacuum filtration. The assay performance was evaluated by real-time qPCR based on Taqman technology. Assay linearity was confirmed with the serial dilutions of RTA (Turkey) containing 1 × (10(6), 10(5), 10(4), and 10(3)) IU mL(−1). Comparison of test results obtained by two extraction methods showed a good correlation (r = 0.95, n = 30) with detection limit of 10(2) IU mL(−1). The semiautomated vacuum filtration based protocol demonstrated high throughput: 35 minutes for the extraction of a batch of 30 samples (150 µL each) with reduced labor, time, waste, and cost. Performance characteristics of semiautomated system make it suitable for use in diagnostic purpose and viral load determinations. Hindawi Publishing Corporation 2015 2015-03-19 /pmc/articles/PMC4383304/ /pubmed/25873968 http://dx.doi.org/10.1155/2015/367801 Text en Copyright © 2015 Imran Amin et al. https://creativecommons.org/licenses/by/3.0/ This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Article
Amin, Imran
Jabbar, Tania
Niazi, Fawad
Akhtar, Muhammad Saeed
Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title_full Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title_fullStr Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title_full_unstemmed Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title_short Extraction of HCV-RNA from Plasma Samples: Development towards Semiautomation
title_sort extraction of hcv-rna from plasma samples: development towards semiautomation
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4383304/
https://www.ncbi.nlm.nih.gov/pubmed/25873968
http://dx.doi.org/10.1155/2015/367801
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