At-home saliva sampling in healthy adults using CandyCollect, a lollipop-inspired device

Tu, Wan-chen; McManamen, Anika M.; Su, Xiaojing; Jeacopello, Ingrid; Takezawa, Meg G.; Hieber, Damielle L.; Hassan, Grant W.; Lee, Ulri N.; Anana, Eden V.; Locknane, Mason P.; Stephenson, Molly W.; Shinkawa, Victoria A. M.; Wald, Ellen R.; DeMuri, Gregory P.; Adams, Karen N.; Thongpang, Sanitta; Theberge, Ashleigh B.

doi:10.1101/2023.01.14.524039

Cited by 2 publications

(1 citation statement)

References 30 publications

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“…In addition to SARS-CoV-2, saliva could serve as specimen for other respiratory viruses [34] such as Influenza viruses, Respiratory Syncytial Virus (RSV), Adenovirus, and Rhinovirus and even bacterial pathogens such as Group A streptococci causing strep throat, and Bordetella pertussis causing whooping cough [35]. This study suggests that the 18S rRNA could be a suitable IAC for other diagnostic tests that rely on saliva samples.…”

Section: Rt-lamp Of 18s Ribosomal Rna Iac Specific To Human Rna In Sa...mentioning

confidence: 99%

Development of an Integrated Sample Amplification Control for Salivary Point-of-Care Pathogen Testing

Sritong,

Ngo,

Ejendal

et al. 2023

Preprint

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BackgroundThe COVID-19 pandemic has led to a rise in point-of-care (POC) and home-based tests, but concerns over usability, accuracy, and effectiveness have arisen. The incorporation of internal amplification controls (IACs), essential control for translational POC diagnostics, could mitigate false-negative and false-positive results due to sample matrix interference or inhibition. Although emerging POC nucleic acid amplification tests (NAATs) for detecting SARS-CoV-2 show impressive analytical sensitivity in the lab, the assessment of clinical accuracy with IACs is often overlooked. In some cases, the IACs were run spatially, complicating assay workflow. Therefore, the multiplex assay for pathogen and IAC is needed.ResultsWe developed a one-pot duplex reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) assay for saliva samples, a non-invasive and simple collected specimen for POC NAATs. The ORF1ab gene of SARS-CoV-2 was used as a target and a human 18S ribosomal RNA in human saliva was employed as an IAC to ensure clinical reliability of the RT-LAMP assay. The optimized assay could detect SARS-CoV-2 viral particles down to 100 copies/μL of saliva within 30 minutes without RNA extraction. The duplex RT-LAMP for SARS-CoV-2 and IAC is successfully amplified in the same reaction without cross-reactivity. The valid results were easily visualized in triple-line lateral flow immunoassay, in which two lines (flow control and IAC lines) represent valid negative results and three lines (flow control, IAC, and test line) represent valid positive results. This duplex assay demonstrated a clinical sensitivity of 95%, specificity of 100%, and accuracy of 96% in 30 clinical saliva samples.SignificanceIACs play a crucial role in ensuring user confidence with respect to the accuracy and reliability of at-home and POC molecular diagnostics. We demonstrated the multiplex capability of SARS-COV-2 and human18S ribosomal RNA RT-LAMP without complicating assay design. This generic platform can be extended in a similar manner to include human18S ribosomal RNA IACs into different clinical sample matrices.

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Section: Rt-lamp Of 18s Ribosomal Rna Iac Specific To Human Rna In Sa...mentioning

confidence: 99%

Development of an Integrated Sample Amplification Control for Salivary Point-of-Care Pathogen Testing

Sritong,

Ngo,

Ejendal

et al. 2023

Preprint

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Integration of Group A Streptococcus Rapid Tests with the Open Fluidic CandyCollect Device

Sanchez,

Robertson,

Shinkawa

et al. 2024

Preprint

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The CandyCollect device is a lollipop-inspired open fluidic oral sampling device designed to provide a comfortable user sampling experience. We demonstrate that the CandyCollect device can be coupled with a rapid antigen detection test (RADT) kit designed for Group A Streptococcus (GAS). Throughin vitroexperiments with pooled saliva spiked withStreptococcus pyogeneswe tested various reagents and elution volumes to optimize the RADT readout from CandyCollect device samples. The resulting optimized protocol uses the kit-provided reagents and lateral flow assay (LFA) while replacing the kit’s pharyngeal swab with the CandyCollect device, reducing the elution solution volume, and substituting the tube used for elution to accommodate the CandyCollect device. Positive test results were detected by eye with bacterial concentrations as low as the manufacturer’s “minimal detection limit” - 1.5×105CFU/mL. LFA strips were also scanned and quantified with image analysis software to determine the signal-to-baseline ratio (SBR) and categorize positive test results without human bias. We tested our optimized protocol for integrating CandyCollect and RADT using CandyCollect clinical samples from pediatric patients (n=6) who were previously diagnosed with GAS pharyngitis via pharyngeal swabs tested with RADT as part of their clinical care. The LFA results of these CandyCollect devices and interspersed negative controls were determined by independent observers, with positive results obtained in four of the six participants on at least one LFA replicate. Taken together, our results show that CandyCollect devices from children with GAS pharyngitis can be tested using LFA rapid tests.Table of Contents/Abstract Figure

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At-home saliva sampling in healthy adults using CandyCollect, a lollipop-inspired device

Cited by 2 publications

References 30 publications

Development of an Integrated Sample Amplification Control for Salivary Point-of-Care Pathogen Testing

Development of an Integrated Sample Amplification Control for Salivary Point-of-Care Pathogen Testing

Integration of Group A Streptococcus Rapid Tests with the Open Fluidic CandyCollect Device

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