Multiplex, Quantitative, Reverse Transcription PCR Detection of Influenza Viruses Using Droplet Microfluidic Technology
نویسندگان
چکیده
Quantitative, reverse transcription, polymerase chain reaction (qRT-PCR) is facilitated by leveraging droplet microfluidic (DMF) system, which due to its precision dispensing and sample handling capabilities at microliter and lower volumes has emerged as a popular method for miniaturization of the PCR platform. This work substantially improves and extends the functional capabilities of our previously demonstrated single qRT-PCR micro-chip, which utilized a combination of electrostatic and electrowetting droplet actuation. In the reported work we illustrate a spatially multiplexed micro-device that is capable of conducting up to eight parallel, real-time PCR reactions per usage, with adjustable control on the PCR thermal cycling parameters (both process time and temperature set-points). This micro-device has been utilized to detect and quantify the presence of two clinically relevant respiratory viruses, Influenza A and Influenza B, in human samples (nasopharyngeal swabs, throat swabs). The device performed accurate detection and quantification of the two respiratory viruses, over several orders of RNA copy counts, in unknown (blind) panels of extracted patient samples with acceptably high PCR efficiency (>94%). The multi-stage qRT-PCR assays on eight panel patient samples OPEN ACCESS Micromachines 2015, 6 64 were accomplished within 35–40 min, with a detection limit for the target Influenza virus RNAs estimated to be less than 10 RNA copies per reaction.
منابع مشابه
Development of Multiplex Reverse Transcription-Polymerase Chain Reaction for Simultaneous Detection of Influenza A, B and Adenoviruses
Background and objective:Millions of people in developing countries lose their lives due to acute respiratory infections, such as Influenza A & B and Adeno viruses. Given the importance of rapid identification of the virus, in this study the researchers attempted to design a method that enables detection of influenza A, B, and adenoviruses, quickly and simultaneously. ...
متن کاملDevelopment and Evaluation of Real-Time Reverse Transcription Polymerase Chain Reaction Test for Quantitative and Qualitative Recognition of H5 Subtype of Avian Influenza Viruses
Avian influenza viruses (AIV) affect a wide range of birds and mammals, cause severe economic damage to the poultry industry, and pose a serious threat to humans. Highly pathogenic avian influenza viruses (HPAI) H5N1 were first identified in Southeast Asia in 1996 and spread to four continents over the following years. The viruses have caused high mortality in chickens and various bird species ...
متن کاملMultiplex Reverse Transcriptase-PCR Assay for Typing and Subtyping of Influenza A (H5 & H9) Virus in Iran
Avian influenza virus (AIV) infection is a major cause of bird or human mortality and morbidity, therefore the rapid identification of the virus is of important clinical and epidemiological implication. Methods: A multiplex Reverse Transcriptase PCR (RT-PCR) was optimized for the detection of influenza A virus and the H5 and H9 subtypes. The influenza type A specific primers were directed to t...
متن کاملDetection of avian influenza virus of H9 subtype in the feces of experimentally infected chickens by RT–PCR
Avian Influenza (AI) is a viral respiratory disease of domestic and wild birds. In the diagnostic laboratory, it is essential to have methods for rapid detection of avian respiratory viruses. Cloacal swabs collected from chickens experimentally infected with H9 subtype AI virus, used in a reverse transcription-polymerase chain reaction (RT-PCR) assay for detection of AI. In infected animals, ...
متن کاملDetection of Seasonal Influenza H1N1 and H3N2 Viruses using RT-PCR Assay during 2009 Flu Pandemic in Golestan Province
Abstract Background and Objective: The emergence of a novel H1N1influenza A virus of animal origin with transmissibility from human to human poses pandemic concern. Current subtypes of Seasonal influenza A viruses spread in human are influenza A H1N1 influenza A H3N2 and influenza type B viruses. The aim of this study was to determine current strains of the H3N2 and new H1N1 subtypes of influe...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Micromachines
دوره 6 شماره
صفحات -
تاریخ انتشار 2015