Molecular test for COVID-19 diagnosis based on a colorimetric genomagnetic assay.

The world is in a long pandemic period caused by the SARS-CoV-2 virus and massive diagnostic tests to assist efforts to control the spread of the disease and also to avoid new coronavirus variants are still needed. Herein, we propose a simple and accurate saliva-based colorimetric test for the diagnosis of COVID-19. Magnetic beads (MBs) modified with a sequence of single-strand DNA (ssDNA) complementary to the N gene of the SARS-CoV-2 RNA were developed and used for magnetic capture and separation from a complex saliva sample. A second biotinylated ssDNA sequence was applied, and the colorimetric detection was carried out by adding streptavidin-horseradish peroxidase conjugate, H2O2, and tetramethylbenzidine (TMB) as chromogenic substrate. The test does not require viral RNA isolation, transcription, or amplification steps and can be performed at room temperature. The molecular assay test can be run using 96-well microplates, allowing the diagnosis of a large number of samples in 90 min. A simple support for magnets was designed and constructed using a 3D printer that allows the magnetic separations directly in the 96-well microplate. The colorimetric test showed an excellent ability to discriminate between healthy individuals and patients infected with SARS-CoV-2, with 92% and 100% of clinical sensitivity and specificity, respectively. This performance was similar to that achieved using the gold standard RT-PCR technique. The proposed genomagnetic assay offers an opportunity to greatly increase population testing, contribute to controlling the spread of the virus, and improve health equity in testing for COVID-19.

Development of RNA-Based Assay for Rapid Detection of SARS-CoV-2 in Clinical Samples.

INTRODUCTION: The ongoing spread of pandemic coronavirus disease-19 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is of growing concern. Rapid diagnosis and management of SARS-CoV-2 are crucial for controlling the outbreak in the community. Here, we report the development of a first rapid-colorimetric assay capable of detecting SARS-CoV-2 in the human nasopharyngeal RNA sample in less than 30 min. METHOD: We utilized a nanomaterial-based optical sensing platform to detect RNA-dependent RNA polymerase gene of SARS-CoV-2, where the formation of oligo probe-target hybrid led to salt-induced aggregation and change in gold-colloid color from pink to blue visibility range. Accordingly, we found a change in colloid color from pink to blue in assay containing nasopharyngeal RNA sample from the subject with clinically diagnosed COVID-19. The colloid retained pink color when the test includes samples from COVID-19 negative subjects or human papillomavirus-infected women. RESULTS: The results were validated using nasopharyngeal RNA samples from positive COVID-19 subjects (n = 136). Using real-time polymerase chain reaction as gold standard, the assay was found to have 85.29% sensitivity and 94.12% specificity. The optimized method has detection limit as little as 0.5 ng of SARS-CoV-2 RNA. CONCLUSION: We found that the developed assay rapidly detects SARS-CoV-2 RNA in clinical samples in a cost-effective manner and would be useful in pandemic management by facilitating mass screening.

Peptide Amphiphile Mediated Co-assembly for Nanoplasmonic Sensing.

Aromatic interactions are commonly involved in the assembly of naturally occurring building blocks, and these interactions can be replicated in an artificial setting to produce functional materials. Here we describe a colorimetric biosensor using co-assembly experiments with plasmonic gold and surfactant-like peptides (SLPs) spanning a wide range of aromatic residues, polar stretches, and interfacial affinities. The SLPs programmed in DDD-(ZZ) x -FFPC self-assemble into higher-order structures in response to a protease and subsequently modulate the colloidal dispersity of gold leading to a colorimetric readout. Results show the strong aggregation propensity of the FFPC tail without polar DDD head. The SLPs were specific to the target protease, i.e., Mpro, a biomarker for SARS-CoV-2. This system is a simple and visual tool that senses Mpro in phosphate buffer, exhaled breath condensate, and saliva with detection limits of 15.7, 20.8, and 26.1 nM, respectively. These results may have value in designing other protease testing methods.