Molecular isothermal techniques for combating infectious diseases: towards low-cost point-of-care diagnostics

Abstract

Nucleic acid amplification techniques such as PCR have facilitated rapid and accurate diagnosis in central laboratories over the past years. PCR-based amplifications require high-precision instruments to perform thermal cycling reactions. Such equipment is bulky, expensive and complex to operate. Progressive advances in isothermal amplification chemistries, microfluidics and detectors miniaturisation are paving the way for the introduction and use of compact 'sample in-results out' diagnostic devices. However, this paradigm shift towards decentralised testing poses diverse technological, economic and organizational challenges both in industrialized and developing countries. This review describes the landscape of molecular isothermal diagnostic techniques for infectious diseases, their characteristics, current state of development, and available products, with a focus on new directions towards point-of-care applications.

Recombinase Polymerase Amplification for Diagnostic Applications

“…In the last decade, there was a remarkable increase in the development and adaptation of novel and existing isothermal amplification technologies for molecular diag-nostics. More than a dozen isothermal technologies, differing in their respective enzymatic mechanisms, are known and have been recently reviewed elsewhere (1)(2)(3). In this review, we focus on recombinase polymerase amplification (RPA) 3 because of its simplicity (few and easy hands-on manipulations), flexibility (different commercial kit formats offering various detection methods for both DNA and RNA), and speed (results in 5-20 min).…”

Laboratory diagnosis of pneumonia in the molecular age

“…Recently, a new group of molecular techniques has emerged based on LAMP [45]. Unlike conventional PCRs, in which a series of temperature change cycles are required for the amplification of nucleic acids, these techniques allow amplification at a constant temperature.…”

Validation of a Loop-Mediated Isothermal Amplification Assay for Rapid Diagnosis of Invasive Pneumococcal Disease

“…In recent years, the emergence of novel isothermal techniques such as nicking endonuclease amplification reaction (NEAR), strand displacement amplification (SDA), loop-mediated isothermal amplification (LAMP), and helicase-dependent amplification (HAD) has led to the emergence of inexpensive and easy-to-use diagnostic devices that fulfill most ASSURED criteria (de Paz et al, 2014). Particularly, the LAMP technique (Notomi, 2000) has been reported to have similar, or even better, diagnostic accuracy than PCR for detecting infectious agents (Chen et al, 2011;Parida et al, 2011;Patel et al, 2014;Brotons et al, 2016), including S. pneumoniae (Seki et al, 2005;Kim et al, 2012).…”

“…In addition, the Bst polymerase has a high tolerance to inhibitory substances commonly present in biological samples. All these features make this molecular chemistry highly suitable for diagnostic use in resource-limited settings (Notomi, 2000;de Paz et al, 2014).…”