Browsing by Author "Iles, Alexander"

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    An integrated lab-on-a-chip device for RNA extraction, amplification and CRISPR-Cas12a-assisteddetection for COVID-19 screening in resource-limited settings
    (MedRXiv, 2022-01-06) Ngamsom, Bongkot; Iles, Alexander; Kamita, Moses; Kimani, Racheal; Rodriguez-Mateos, Pablo; Mungai, Mary; Dyer,Charlotte; Walter, Cheryl; Pamme, Nicole; Gitaka, Jesse
    n response to the ongoing COVID-19 pandemic and disparities of vaccination coverage in low-and middle-incomecountries, it is vital to adopt a widespread testing and screening programme, combined with contact tracing, to monitor and effectively control the infection dispersion in areas where medical resources are limited.This work presents a lab-on-a-chip platform, namely“IFAST-CRISPR”, as an affordable, rapid and high-precision molecular diagnostic means for SARS-CoV-2 detection. The herein proposed “sample-to-answer”platform integratesRNA extraction, amplification and CRISPR-Cas-based detection with lateral flow readout in one device. The microscale dimensions of the device containing immiscible liquids, coupled with the use of silica paramagnetic beads and GuHCl, streamline sample preparation, including RNA concentration, extraction and purification, in15 min with minimal hands-on steps. By combining RT-LAMP with CRISPR-Cas12 assays targeting the nucleoprotein (N) gene, visual identification of ≥470 copies mL-1genomic SARS-CoV-2 sampleswas achieved in 45 min, with no cross-reactivity towards HCoV-OC43 nor H1N1. On-chip assays showed the ability to isolate and detect SARS-CoV-2 from 1,000 genome copies mL-1of replication-deficient viral particles in 1 h. This simple, affordable and integrated platform demonstrated a visual, faster, and yet specificity and sensitivity-comparable alternative to the costly gold-standard RT-PCR assay, requiring only a simple heating source. Further investigations on multiplexing and direct interfacing of the accessible Swan-brand cigarette filter for saliva sample collectioncould providea complete work flow for COVID-19 diagnostics from saliva samples suitable for low-resource settings.
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    Rapid detection of Group B Streptococcus (GBS) from artificial urine samples based on IFAST and ATP bioluminescence assay: from development to practical challenges during protocol testing in Kenya
    (Analyst, 2019-10-10) Ngamsom, Bongkot; Wandera, Ernest Apondi; Iles, Alexander; Kimani, Racheal; Muregi, Francis; Gitaka, Jesse; Pamme, Nicole
    We report the rapid detection (20 min) of Streptococcus agalactiae, Group B Streptococcus (GBS) employing on-chip magnetic isolation of GBS based on immiscible filtration assisted by surface tension (IFAST), followed by detection of the isolated GBS using an adenosine triphosphate (ATP) bioluminescence assay. Up to 80% GBS cells were isolated from spiked artificial urine samples with linear responses of bioluminescence signals from isolated cells at 2.3 × 102–9.1 × 105 CFU mL−1, demonstrating great promise for point-of-care detection of pathogenic bacteria in screening urine samples from pregnant women. Practical challenges during initial testing of the developed protocol with urine samples in Kenya are also described.
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    Rapid detection of Group B Streptococcus (GBS) from artificial urine samples based on IFAST and ATP bioluminescence assay: from development to practical challenges during protocol testing in Kenya
    (Royal Society of Chemistry, 2019-10-10) Pamme, Nicole; Wandera,Ernest Apondi; Ngamsom, Bongkot; Iles, Alexander; Kimani, Racheal; Muregi,Francis; Gitaka, Jesse
    We report the rapid detection (20 min) of Streptococcus agalactiae, Group B Streptococcus (GBS) employing on-chip magnetic isolation of GBS based on immiscible filtration assisted by surface tension (IFAST), followed by detection of the isolated GBS using an adenosine triphosphate (ATP) bioluminescence assay. Up to 80% GBS cells were isolated from spiked artificial urine samples with linear responses of bioluminescence signals from isolated cells at 2.3 × 102–9.1 × 105 CFU mL−1, demonstrating great promise for point-of-care detection of pathogenic bacteria in screening urine samples from pregnant women. Practical challenges during initial testing of the developed protocol with urine samples in Kenya are also described.