Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study

Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we...
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Autor*in:	Chiu, Rossa WK [verfasserIn] Jin, Yongjie Chung, Grace TY Lui, Wing-bong Chan, Anthony TC Lim, Wilina Dennis Lo, YM

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2006

Schlagwörter:	Reverse Transcriptase Polymerase Chain Reaction Severe Acute Respiratory Syndrome Total Nucleic Acid Reverse Transcriptase Polymerase Chain Reaction Assay

Anmerkung:	© Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (

Übergeordnetes Werk:	Enthalten in: BMC infectious diseases - London : BioMed Central, 2001, 6(2006), 1 vom: 09. Feb.
Übergeordnetes Werk:	volume:6 ; year:2006 ; number:1 ; day:09 ; month:02

Links:	Volltext

DOI / URN:	10.1186/1471-2334-6-20

Katalog-ID:	SPR027692310

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520			\|a Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method.
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700	1		\|a Jin, Yongjie \|4 aut
700	1		\|a Chung, Grace TY \|4 aut
700	1		\|a Lui, Wing-bong \|4 aut
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700	1		\|a Lim, Wilina \|4 aut
700	1		\|a Dennis Lo, YM \|4 aut
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allfields	10.1186/1471-2334-6-20 doi (DE-627)SPR027692310 (SPR)1471-2334-6-20-e DE-627 ger DE-627 rakwb eng Chiu, Rossa WK verfasserin aut Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study 2006 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213 Jin, Yongjie aut Chung, Grace TY aut Lui, Wing-bong aut Chan, Anthony TC aut Lim, Wilina aut Dennis Lo, YM aut Enthalten in BMC infectious diseases London : BioMed Central, 2001 6(2006), 1 vom: 09. Feb. (DE-627)326645381 (DE-600)2041550-3 1471-2334 nnns volume:6 year:2006 number:1 day:09 month:02 https://dx.doi.org/10.1186/1471-2334-6-20 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_702 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2031 GBV_ILN_2038 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2056 GBV_ILN_2057 GBV_ILN_2061 GBV_ILN_2111 GBV_ILN_2113 GBV_ILN_2190 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 6 2006 1 09 02
spelling	10.1186/1471-2334-6-20 doi (DE-627)SPR027692310 (SPR)1471-2334-6-20-e DE-627 ger DE-627 rakwb eng Chiu, Rossa WK verfasserin aut Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study 2006 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213 Jin, Yongjie aut Chung, Grace TY aut Lui, Wing-bong aut Chan, Anthony TC aut Lim, Wilina aut Dennis Lo, YM aut Enthalten in BMC infectious diseases London : BioMed Central, 2001 6(2006), 1 vom: 09. Feb. (DE-627)326645381 (DE-600)2041550-3 1471-2334 nnns volume:6 year:2006 number:1 day:09 month:02 https://dx.doi.org/10.1186/1471-2334-6-20 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_702 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2031 GBV_ILN_2038 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2056 GBV_ILN_2057 GBV_ILN_2061 GBV_ILN_2111 GBV_ILN_2113 GBV_ILN_2190 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 6 2006 1 09 02
allfields_unstemmed	10.1186/1471-2334-6-20 doi (DE-627)SPR027692310 (SPR)1471-2334-6-20-e DE-627 ger DE-627 rakwb eng Chiu, Rossa WK verfasserin aut Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study 2006 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213 Jin, Yongjie aut Chung, Grace TY aut Lui, Wing-bong aut Chan, Anthony TC aut Lim, Wilina aut Dennis Lo, YM aut Enthalten in BMC infectious diseases London : BioMed Central, 2001 6(2006), 1 vom: 09. Feb. (DE-627)326645381 (DE-600)2041550-3 1471-2334 nnns volume:6 year:2006 number:1 day:09 month:02 https://dx.doi.org/10.1186/1471-2334-6-20 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_702 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2031 GBV_ILN_2038 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2056 GBV_ILN_2057 GBV_ILN_2061 GBV_ILN_2111 GBV_ILN_2113 GBV_ILN_2190 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 6 2006 1 09 02
allfieldsGer	10.1186/1471-2334-6-20 doi (DE-627)SPR027692310 (SPR)1471-2334-6-20-e DE-627 ger DE-627 rakwb eng Chiu, Rossa WK verfasserin aut Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study 2006 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213 Jin, Yongjie aut Chung, Grace TY aut Lui, Wing-bong aut Chan, Anthony TC aut Lim, Wilina aut Dennis Lo, YM aut Enthalten in BMC infectious diseases London : BioMed Central, 2001 6(2006), 1 vom: 09. Feb. (DE-627)326645381 (DE-600)2041550-3 1471-2334 nnns volume:6 year:2006 number:1 day:09 month:02 https://dx.doi.org/10.1186/1471-2334-6-20 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_702 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2031 GBV_ILN_2038 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2056 GBV_ILN_2057 GBV_ILN_2061 GBV_ILN_2111 GBV_ILN_2113 GBV_ILN_2190 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 6 2006 1 09 02
allfieldsSound	10.1186/1471-2334-6-20 doi (DE-627)SPR027692310 (SPR)1471-2334-6-20-e DE-627 ger DE-627 rakwb eng Chiu, Rossa WK verfasserin aut Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study 2006 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213 Jin, Yongjie aut Chung, Grace TY aut Lui, Wing-bong aut Chan, Anthony TC aut Lim, Wilina aut Dennis Lo, YM aut Enthalten in BMC infectious diseases London : BioMed Central, 2001 6(2006), 1 vom: 09. Feb. (DE-627)326645381 (DE-600)2041550-3 1471-2334 nnns volume:6 year:2006 number:1 day:09 month:02 https://dx.doi.org/10.1186/1471-2334-6-20 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_702 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2031 GBV_ILN_2038 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2056 GBV_ILN_2057 GBV_ILN_2061 GBV_ILN_2111 GBV_ILN_2113 GBV_ILN_2190 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 6 2006 1 09 02
language	English
source	Enthalten in BMC infectious diseases 6(2006), 1 vom: 09. Feb. volume:6 year:2006 number:1 day:09 month:02
sourceStr	Enthalten in BMC infectious diseases 6(2006), 1 vom: 09. Feb. volume:6 year:2006 number:1 day:09 month:02
format_phy_str_mv	Article
institution	findex.gbv.de
topic_facet	Reverse Transcriptase Polymerase Chain Reaction Severe Acute Respiratory Syndrome Total Nucleic Acid Reverse Transcriptase Polymerase Chain Reaction Assay
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container_title	BMC infectious diseases
authorswithroles_txt_mv	Chiu, Rossa WK @@aut@@ Jin, Yongjie @@aut@@ Chung, Grace TY @@aut@@ Lui, Wing-bong @@aut@@ Chan, Anthony TC @@aut@@ Lim, Wilina @@aut@@ Dennis Lo, YM @@aut@@
publishDateDaySort_date	2006-02-09T00:00:00Z
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author	Chiu, Rossa WK
spellingShingle	Chiu, Rossa WK misc Reverse Transcriptase Polymerase Chain Reaction misc Severe Acute Respiratory Syndrome misc Total Nucleic Acid misc Reverse Transcriptase Polymerase Chain Reaction Assay Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study
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topic_title	Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study Reverse Transcriptase Polymerase Chain Reaction (dpeaa)DE-He213 Severe Acute Respiratory Syndrome (dpeaa)DE-He213 Total Nucleic Acid (dpeaa)DE-He213 Reverse Transcriptase Polymerase Chain Reaction Assay (dpeaa)DE-He213
topic	misc Reverse Transcriptase Polymerase Chain Reaction misc Severe Acute Respiratory Syndrome misc Total Nucleic Acid misc Reverse Transcriptase Polymerase Chain Reaction Assay
topic_unstemmed	misc Reverse Transcriptase Polymerase Chain Reaction misc Severe Acute Respiratory Syndrome misc Total Nucleic Acid misc Reverse Transcriptase Polymerase Chain Reaction Assay
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title	Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study
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title_full	Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study
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author_browse	Chiu, Rossa WK Jin, Yongjie Chung, Grace TY Lui, Wing-bong Chan, Anthony TC Lim, Wilina Dennis Lo, YM
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title_sort	automated extraction protocol for quantification of sars-coronavirus rna in serum: an evaluation study
title_auth	Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study
abstract	Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (
abstractGer	Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (
abstract_unstemmed	Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method. © Chiu et al; licensee BioMed Central Ltd. 2006. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (
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title_short	Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study
url	https://dx.doi.org/10.1186/1471-2334-6-20
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author2	Jin, Yongjie Chung, Grace TY Lui, Wing-bong Chan, Anthony TC Lim, Wilina Dennis Lo, YM
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Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study

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