Figure 6 Sensitivity and specificity of Colony PCR reactions. L: DNA ladder, N: Negative sample, P: Positive sample, R: Colony diameter. E. coli: Rmin ≈ 1.5 mm, Rmean ≈ 1.8 mm, Rmax ≈ 2.5 mm. P. aeruginosa : R ≈ 2 mm.
Here, for the first time we presented a Colony-LAMP assay performed in the LAMP device that takes the samples and provides real-time colorimetric readouts in 30 minutes. While an early work by Yan and co-workers implemented a Colony-LAMP assay for 116 strains of food pathogens using analytical laboratory setups, this is the first work that developed a LAMP device capable of conducting both conventional and Colony-LAMP reactions [14]. Besides, this work is one of the first examples of field-ready modern LAMP device that is low-cost, portable, user-friendly, and robust to rapidly provide specific and sensitive readouts without false positives. In our previous studies, we developed a simple, low-cost, and portable LAMP device for GMO detection in the field [10]. Next, similar technologies have been developed to perform LAMP reactions either for COVID-19 diagnosis [23, 26-28] or food pathogen detection [24, 25, 29-31]. Still, there is a pressing need for a sensitive, robust, and reliable integrated method of nucleic acid extraction and detection on the same diagnostic devices. In other words, nucleic acid-based diagnostic technologies should be capable of utilizing the entire process from sample preparation to visualization of the results at once.
The major hurdles for on-site nucleic acid detection can be classified as method- and technology-based limitations. Most of the nucleic acid detection methodologies involves nucleic acid extraction and amplification steps requiring tedious liquid handling steps with high-cost reagents, specialized equipment, and trained technicians [1-8]. These hurdles limit their usefulness for field-based studies. LAMP technique has overcome majority of these limitations [2]. Improved and more sample-specific LAMP protocols have also been developed and published in recent decade [9-19]. In contrast to LAMP methods, technology development for LAMP-based detection has not been completely accelerated yet. In most studies, the focus is developing microfluidic devices for LAMP-based assays. However, microfluidic devices are expensive to fabricate, complex to use, and require trained personnel and stable operation conditions, which makes this technology not feasible for field deployable real-time diagnosis [30-33]. Therefore, portable, rapid, cost-effective, automated, and remotely controllable LAMP platforms with colorimetric readouts will be future of field-ready modern nucleic-acid based detection platforms.