Polyethersulfone improves isothermal nucleic acid amplification compared to current paper-based diagnostics

Abstract

Devices based on rapid, paper-based, isothermal nucleic acid amplification techniques have recently emerged with the potential to fill a growing need for highly sensitive point-of-care diagnostics throughout the world. As this field develops, such devices will require optimized materials that promote amplification and sample preparation. Herein, we systematically investigated isothermal nucleic acid amplification in materials currently used in rapid diagnostics (cellulose paper, glass fiber, and nitrocellulose) and two additional porous membranes with upstream sample preparation capabilities (polyethersulfone and polycarbonate). We compared amplification efficiency from four separate DNA and RNA targets (Bordetella pertussis, Chlamydia trachomatis, Neisseria gonorrhoeae, and Influenza A H1N1) within these materials using two different isothermal amplification schemes, helicase dependent amplification (tHDA) and loop-mediated isothermal amplification (LAMP), and traditional PCR. We found that the current paper-based diagnostic membranes inhibited nucleic acid amplification when compared to membrane-free controls; however, polyethersulfone allowed for efficient amplification in both LAMP and tHDA reactions. Further, observing the performance of traditional PCR amplification within these membranes was not predicative of their effects on in situ LAMP and tHDA. Polyethersulfone is a new material for paper-based nucleic acid amplification, yet provides an optimal support for rapid molecular diagnostics for point-of-care applications.

A paperfluidic platform to detect Neisseria gonorrhoeae in clinical samples

“…Previously confirmed as PES compatible (Linnes et al 2016), our tHDA assay amplifies the highly conserved and single-copy porA pseudogene, which is specific to the NG genome (Unemo et al 2005). Like any molecular target, mutations of the porA gene could result in false-negative results test results.…”

Paper microfluidics for nucleic acid amplification testing (NAAT) of infectious diseases

“…Notwithstanding this, the intensities of the test and control lines were analyzed and measured using ImageJ (50), and the intensity of the control line (in the red channel) was divided by the intensity of the test line for each strip to obtain the percentage intensity of the control line for each test (51).…”

<p>Current and Future Perspectives on Isothermal Nucleic Acid Amplification Technologies for Diagnosing Infections</p>

“…Polyethersulfone though not widely applied in rapid diagnostic assays, proved to be a better material for rapid POC application with nucleic acid from Bordetella pertussis, Chlamydia trachomatis, Neisseria gonorrhoeae, and Influenza A H1N1. 167 One major challenge of microfluidic devices is that of biomolecular interactions with the walls of microfluidic channels, which can reduce enzyme and primer concentrations, thereby resulting in reaction inhibition. However, the use of polyethersulfone may be more expensive for application than cellulose.…”