Giuseppe
Emeritus
[Source: PLoS ONE, full text: (LINK). Abstract, edited.]
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Performance Evaluation of the Maxwell 16 System for Extraction of Influenza Virus RNA from Diverse Samples
Hongbo Liu<SUP>1</SUP><SUP>,</SUP><SUP>2</SUP>, Yan Gan<SUP>3</SUP>, Bo Yang<SUP>4</SUP>, Hui Weng<SUP>1</SUP>, Chunmei Huang<SUP>1</SUP>, Daofeng Yang<SUP>5</SUP>, Ping Lei<SUP>1</SUP>, Guanxin Shen<SUP>1</SUP><SUP>*</SUP>
<SUP></SUP>
1 Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People?s Republic of China, 2 Virology Laboratory, Zhongshan Center for Disease Control and Prevention, Zhongshan, People?s Republic of China, 3 Neuroimmunology Laboratory, Barrow Neurological Institute, St. Joseph?s Hospital and Medical Center, Phoenix, Arizona, United States of America, 4 Department of Intensive Care Unit, Boai Hospital of Zhongshan, Zhongshan, People?s Republic of China, 5 Department of Infectious Diseases, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, People?s Republic of China
Abstract
This study evaluated the performance of the Maxwell 16 System (Promega) for extraction of influenza virus (flu-v) RNA from diverse samples compared to a classical manual method (QIAamp Kit, QIAGEN). Following extraction by the two methods, all samples were analyzed by Real-time RT-PCR. Results revealed that the use of the standard Maxwell 16 protocol (Maxwell 16-S) resulted in good linearity and precision across a wide concentration range and higher sensitivity of detection from flu-v stock suspensions than the manual method. Compared with the latter method, Maxwell 16-S extracted RNA more efficiently (higher RNA yield and/or fewer PCR inhibitors) from throat swabs and bronchoalveolar lavage fluids, while both methods performed comparably on fecal samples from human and poultry in terms of overall threshold cycle values and detection rates although the Maxwell 16-S co-purified more inhibitors from fecal samples. The capacity of this system to remove inhibitors from fecal matrix was improved by using a modified Maxwell 16 protocol with a reduced sample input, which eliminated all false-negatives produced by the Maxwell 16-S. These findings suggest that the Maxwell 16 System is suitable for RNA extraction from multiple-source samples for diagnosis of influenza and viral load determination and that a proper reduction in starting sample volume may improve the detection of flu-v from complex matrices such as feces. Additionally, this system allows flexible sample throughput and labor-saving sample processing with little or no risk of cross-contamination.
Citation: Liu H, Gan Y, Yang B, Weng H, Huang C, et al. (2012) Performance Evaluation of the Maxwell 16 System for Extraction of Influenza Virus RNA from Diverse Samples. PLoS ONE 7(10): e48094. doi:10.1371/journal.pone.0048094
Editor: Suryaprakash Sambhara, Centers for Disease Control and Prevention, United States of America
Received: June 4, 2012; Accepted: September 19, 2012; Published: October 29, 2012
Copyright: ? 2012 Liu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This research work was supported by Program for Changjiang Scholars and Innovative Research Team in University (IRT1131) and State Project on Major Infectious Diseases Prevention (2012ZX10002006-003), the International Science & Technology Cooperation Program of China (2011DFA31030) and Deutsche Forschungsgemeinschaft (SFB/Transregio TRR60). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
* E-mail: guanxin_shen@yahoo.com.cn
-Hongbo Liu<SUP>1</SUP><SUP>,</SUP><SUP>2</SUP>, Yan Gan<SUP>3</SUP>, Bo Yang<SUP>4</SUP>, Hui Weng<SUP>1</SUP>, Chunmei Huang<SUP>1</SUP>, Daofeng Yang<SUP>5</SUP>, Ping Lei<SUP>1</SUP>, Guanxin Shen<SUP>1</SUP><SUP>*</SUP>
<SUP></SUP>
1 Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People?s Republic of China, 2 Virology Laboratory, Zhongshan Center for Disease Control and Prevention, Zhongshan, People?s Republic of China, 3 Neuroimmunology Laboratory, Barrow Neurological Institute, St. Joseph?s Hospital and Medical Center, Phoenix, Arizona, United States of America, 4 Department of Intensive Care Unit, Boai Hospital of Zhongshan, Zhongshan, People?s Republic of China, 5 Department of Infectious Diseases, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, People?s Republic of China
Abstract
This study evaluated the performance of the Maxwell 16 System (Promega) for extraction of influenza virus (flu-v) RNA from diverse samples compared to a classical manual method (QIAamp Kit, QIAGEN). Following extraction by the two methods, all samples were analyzed by Real-time RT-PCR. Results revealed that the use of the standard Maxwell 16 protocol (Maxwell 16-S) resulted in good linearity and precision across a wide concentration range and higher sensitivity of detection from flu-v stock suspensions than the manual method. Compared with the latter method, Maxwell 16-S extracted RNA more efficiently (higher RNA yield and/or fewer PCR inhibitors) from throat swabs and bronchoalveolar lavage fluids, while both methods performed comparably on fecal samples from human and poultry in terms of overall threshold cycle values and detection rates although the Maxwell 16-S co-purified more inhibitors from fecal samples. The capacity of this system to remove inhibitors from fecal matrix was improved by using a modified Maxwell 16 protocol with a reduced sample input, which eliminated all false-negatives produced by the Maxwell 16-S. These findings suggest that the Maxwell 16 System is suitable for RNA extraction from multiple-source samples for diagnosis of influenza and viral load determination and that a proper reduction in starting sample volume may improve the detection of flu-v from complex matrices such as feces. Additionally, this system allows flexible sample throughput and labor-saving sample processing with little or no risk of cross-contamination.
Citation: Liu H, Gan Y, Yang B, Weng H, Huang C, et al. (2012) Performance Evaluation of the Maxwell 16 System for Extraction of Influenza Virus RNA from Diverse Samples. PLoS ONE 7(10): e48094. doi:10.1371/journal.pone.0048094
Editor: Suryaprakash Sambhara, Centers for Disease Control and Prevention, United States of America
Received: June 4, 2012; Accepted: September 19, 2012; Published: October 29, 2012
Copyright: ? 2012 Liu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This research work was supported by Program for Changjiang Scholars and Innovative Research Team in University (IRT1131) and State Project on Major Infectious Diseases Prevention (2012ZX10002006-003), the International Science & Technology Cooperation Program of China (2011DFA31030) and Deutsche Forschungsgemeinschaft (SFB/Transregio TRR60). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
* E-mail: guanxin_shen@yahoo.com.cn
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