Integration of reverse transcriptase loop-mediated isothermal amplification with an immunochromatographic strip on a centrifugal microdevice for influenza A virus identification

Abstract

A novel centrifugal microdevice which could perform reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) and immunochromatographic strip (ICS) based amplicon detection was demonstrated for simple and cost-effective influenza A virus identification. The proposed centrifugal microdevice consists of the sample and running buffer loading reservoirs, the RT-LAMP chamber, and the ICS for detecting gene expression. The entire process could be completed sequentially and automatically by simply controlling the rotation speed and by optimizing the microfluidic design. Monoplex and multiplex RT-LAMP reactions targeting H1 and/or M gene were executed at 66 °C for 40 min, and the resultant amplicons were successfully analysed on the ICS within 15 min. Influenza A H1N1 virus was subtyped by detecting H1 and M gene on the ICS even with 10 copies of viral RNAs. Highly specific and multiplex viral typing of the integrated RT-LAMP-ICS microdevice was also demonstrated. The combination of the rapid isothermal amplification with the simple colorimetric detection on a strip in a single centrifugal microdevice will provide an advanced genetic analysis platform in the field of on-site pathogen diagnostics.

Emerging Loop-Mediated Isothermal Amplification-Based Microchip and Microdevice Technologies for Nucleic Acid Detection

“…54 Each microfluidic device consisted of an inlet for injecting LAMP reagents, a chamber for amplification, and a zigzag-shaped dispensing microchannel which was set to aliquot LAMP solution equally in three chambers. A 2 μ L sample was injected into the inlet and filled the zigzag channel by capillary action.…”

“…(E) Actual image of the assay results for detecting (i) H1 gene, (ii) M gene, and (iii) negative control. Reproduced with permission from ref 54. Copyright 2015 Royal Society of Chemistry.…”

On Facing the SARS-CoV-2 (COVID-19) with Combination of Nanomaterials and Medicine: Possible Strategies and First Challenges

“…These limitations can be resolved by the implementation of nanoparticles (NPs) based therapeutic approaches due to their features such as size and shape control and surface functionalization which ultimately leads to strong immunogenicity and enhanced antigen presentation. Recent literature suggests the toxicity concerns of NPs as quantum dots which need to be carefully addressed while designing theranostic NPs for respiratory diseases [79].…”

LAMP-on-a-chip: Revising microfluidic platforms for loop-mediated DNA amplification

“…Since then, it has attracted a lot of attention and has become an increasingly popular method for clinical diagnosis. So far, LAMP has been employed for rapid detection of both DNA and RNA possessing human-and nonhuman-carried viruses, such as MRSA [43], pseudorabies [24], tuberculosis [72], influenza A [21], Zika [48], nervous necrosis [47], and aqua pathogens in fish [81].…”