A novel Real Time micro PCR based Point-of-Care device for Salmonella detection in human clinical samples

2012 ◽  
Vol 32 (1) ◽  
pp. 259-265 ◽  
Author(s):  
Dolores Verdoy ◽  
Ziortza Barrenetxea ◽  
Javier Berganzo ◽  
Maria Agirregabiria ◽  
Jesús M. Ruano-López ◽  
...  
Keyword(s):  
Real Time ◽  
Point Of Care ◽  
Clinical Samples

scholarly journals Evaluation of mobile real-time polymerase chain reaction tests for the detection of severe acute respiratory syndrome coronavirus 2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chukwunonso Onyilagha ◽  
Henna Mistry ◽  
Peter Marszal ◽  
Mathieu Pinette ◽  
Darwyn Kobasa ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Real Time ◽  
Point Of Care ◽  
Middle Eastern ◽  
Turnaround Time ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Diarrhea Virus ◽  
Highly Sensitive ◽  
Transmissible Gastroenteritis

AbstractThe coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), calls for prompt and accurate diagnosis and rapid turnaround time for test results to limit transmission. Here, we evaluated two independent molecular assays, the Biomeme SARS-CoV-2 test, and the Precision Biomonitoring TripleLock SARS-CoV-2 test on a field-deployable point-of-care real-time PCR instrument, Franklin three9, in combination with Biomeme M1 Sample Prep Cartridge Kit for RNA 2.0 (M1) manual extraction system for rapid, specific, and sensitive detection of SARS-COV-2 in cell culture, human, and animal clinical samples. The Biomeme SARS-CoV-2 assay, which simultaneously detects two viral targets, the orf1ab and S genes, and the Precision Biomonitoring TripleLock SARS-CoV-2 assay that targets the 5′ untranslated region (5′ UTR) and the envelope (E) gene of SARS-CoV-2 were highly sensitive and detected as low as 15 SARS-CoV-2 genome copies per reaction. In addition, the two assays were specific and showed no cross-reactivity with Middle Eastern respiratory syndrome coronavirus (MERS-CoV), infectious bronchitis virus (IBV), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis (TGE) virus, and other common human respiratory viruses and bacterial pathogens. Also, both assays were highly reproducible across different operators and instruments. When used to test animal samples, both assays equally detected SARS-CoV-2 genetic materials in the swabs from SARS-CoV-2-infected hamsters. The M1 lysis buffer completely inactivated SARS-CoV-2 within 10 min at room temperature enabling safe handling of clinical samples. Collectively, these results show that the Biomeme and Precision Biomonitoring TripleLock SARS-CoV-2 mobile testing platforms could reliably and promptly detect SARS-CoV-2 in both human and animal clinical samples in approximately an hour and can be used in remote areas or health care settings not traditionally serviced by a microbiology laboratory.

scholarly journals Toward point-of-care diagnostics of Candida auris

2020 ◽  
Author(s):  
Geoffrey Mulberry ◽  
Sudha Chaturvedi ◽  
Vishnu Chaturvedi ◽  
Brian N. Kim
Keyword(s):  
New York ◽  
Real Time ◽  
Real Time Pcr ◽  
Positive Impact ◽  
Point Of Care ◽  
Low Cost ◽  
High Sensitivity ◽  
Clinical Samples ◽  
Pcr Assay ◽  
Candida Auris

AbstractCandida auris is a multidrug-resistant yeast that presents global health threat for the hospitalized patients. Early diagnostic of C. auris is crucial in control, prevention, and treatment. Candida auris is difficult to identify with standard laboratory methods and often can be misidentified leading to inappropriate management. A newly-devised real-time PCR assay played an important role in the ongoing investigation of the C. auris outbreak in New York metropolitan area. The assay can rapidly detect C. auris DNA in surveillance and clinical samples with high sensitivity and specificity, and also useful for confirmation of C. auris cultures. Despite its positive impact, the real-time PCR assay is difficult to deploy at frontline laboratories due to high-complexity set-up and operation. Using a low-cost handheld real-time PCR device, we show that the C. auris can potentially be identified in a low-complexity assay without the need for high-cost equipment. An implementation of low-cost real-time PCR device in hospitals and healthcare facilities is likely to accelerate the diagnosis of C. auris and for control of the global epidemic.

scholarly journals A handheld point-of-care system for rapid detection of SARS-CoV-2 in under 20 minutes

2020 ◽  
Author(s):  
Jesus Rodriguez-Manzano ◽  
Kenny Malpartida-Cardenas ◽  
Nicolas Moser ◽  
Ivana Pennisi ◽  
Matthew Cavuto ◽  
...  
Keyword(s):  
Real Time ◽  
Limit Of Detection ◽  
Point Of Care ◽  
Lamp Assay ◽  
High Rate ◽  
Epidemiological Surveillance ◽  
Clinical Samples ◽  
Lab On Chip ◽  
Point Of Care System ◽  
On Chip

AbstractThe COVID-19 pandemic is a global health emergency characterized by the high rate of transmission and ongoing increase of cases globally. Rapid point-of-care (PoC) diagnostics to detect the causative virus, SARS-CoV-2, are urgently needed to identify and isolate patients, contain its spread and guide clinical management. In this work, we report the development of a rapid PoC diagnostic test (< 20 min) based on reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) and semiconductor technology for the detection of SARS-CoV-2 from extracted RNA samples. The developed LAMP assay was tested on a real-time benchtop instrument (RT-qLAMP) showing a lower limit of detection of 10 RNA copies per reaction. It was validated against 183 clinical samples including 127 positive samples (screened by the CDC RT-qPCR assay). Results showed 90.55% sensitivity and 100% specificity when compared to RT-qPCR and average positive detection times of 15.45 ± 4.43 min. For validating the incorporation of the RT-LAMP assay onto our PoC platform (RT-eLAMP), a subset of samples was tested (n=40), showing average detection times of 12.89 ± 2.59 min for positive samples (n=34), demonstrating a comparable performance to a benchtop commercial instrument. Paired with a smartphone for results visualization and geo-localization, this portable diagnostic platform with secure cloud connectivity will enable real-time case identification and epidemiological surveillance.One Sentence SummaryWe demonstrate isothermal detection of SARS-CoV-2 in under 20 minutes from extracted RNA samples with a handheld Lab-on-Chip platform.

scholarly journals Real-time fluorometric and end-point colorimetric isothermal assays for detection of equine pathogens C. psittaci and equine herpes virus 1: validation, comparison and application at the point of care

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Martina Jelocnik ◽  
Sharon Nyari ◽  
Susan Anstey ◽  
Nicole Playford ◽  
Tamieka A. Fraser ◽  
...  
Keyword(s):  
Real Time ◽  
Real World ◽  
Point Of Care ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
Analytical Sensitivity ◽  
Equine Herpesvirus 1 ◽  
Diagnostic Assays ◽  
End Point ◽  
Point Detection

Abstract Background C. psittaci has recently emerged as an equine abortigenic pathogen causing significant losses to the Australian Thoroughbred industry, while Equine herpesvirus-1 (EHV-1) is a well-recognized abortigenic agent. Diagnosis of these agents is based on molecular assays in diagnostic laboratories. In this study, we validated C. psittaci and newly developed EHV-1 Loop Mediated Isothermal Amplification (LAMP) assays performed in a real-time fluorometer (rtLAMP) against the reference diagnostic assays. We also evaluated isothermal amplification using commercially available colorimetric mix (cLAMP), and SYBR Green DNA binding dye (sgLAMP) for “naked eye” end-point detection when testing ‘real-world’ clinical samples. Finally, we applied the C. psittaci LAMP assays in two pilot Point-of-Care (POC) studies in an equine hospital. Results The analytical sensitivity of C. psittaci and EHV-1 rt-, and colorimetric LAMPs was determined as one and 10 genome equivalents per reaction, respectively. Compared to reference diagnostic qPCR assays, the C. psittaci rtLAMP showed sensitivity of 100%, specificity of 97.5, and 98.86% agreement, while EHV-1 rtLAMP showed 86.96% sensitivity, 100% specificity, and 91.43% agreement. When testing rapidly processed clinical samples, all three C. psittaci rt-, c-, sg-LAMP assays were highly congruent with each other, with Kappa values of 0. 906 for sgLAMP and 0. 821 for cLAMP when compared to rtLAMP. EHV-1 testing also revealed high congruence between the assays, with Kappa values of 0.784 for cLAMP and 0.638 for sgLAMP when compared to rtLAMP. The congruence between LAMP assays and the C. psittaci or EHV-1 qPCR assays was high, with agreements ranging from 94.12 to 100% for C. psittaci, and 88.24 to 94.12% for EHV-1, respectively. At the POC, the C. psittaci rt- and c-LAMP assays using rapidly processed swabs were performed by technicians with no prior molecular experience, and the overall congruence between the POC C. psittaci LAMPs and the qPCR assays ranged between 90.91–100%. Conclusions This study describes reliable POC options for the detection of the equine pathogens: C. psittaci and EHV-1. Testing ‘real-world’ samples in equine clinical setting, represents a proof-of-concept that POC isothermal diagnostics can be applied to rapid disease screening in the equine industry.

Erratum to “A novel Real Time micro PCR based Point-of-Care device for Salmonella detection in human clinical samples” [Biosens. Bioelectron. 32 (2012) 259–265]

2012 ◽  
Vol 33 (1) ◽  
pp. 314
Author(s):  
Dolores Verdoy ◽  
Ziortza Barrenetxea ◽  
Javier Berganzo ◽  
Maria Agirregabiria ◽  
Jesús M. Ruano-López ◽  
...  
Keyword(s):  
Real Time ◽  
Point Of Care ◽  
Clinical Samples

scholarly journals Development and evaluation of rapid novel isothermal amplification assays for important veterinary pathogens:Chlamydia psittaciandChlamydia pecorum

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3799 ◽  
Author(s):  
Martina Jelocnik ◽  
Md. Mominul Islam ◽  
Danielle Madden ◽  
Cheryl Jenkins ◽  
James Branley ◽  
...  
Keyword(s):  
Real Time ◽  
Pathogen Detection ◽  
Point Of Care ◽  
Chlamydia Psittaci ◽  
Isothermal Amplification ◽  
Diagnostic Tools ◽  
Clinical Samples ◽  
Conserved Gene ◽  
Thermal Cycler ◽  
Species Specific

BackgroundChlamydia psittaciandChlamydia pecorumare important veterinary pathogens, with the former also being responsible for zoonoses, and the latter adversely affecting koala populations in Australia and livestock globally. The rapid detection of these organisms is still challenging, particularly at the point-of-care (POC). In the present study, we developed and evaluated rapid, sensitive and robustC. psittaci-specific andC. pecorum-specific Loop Mediated Isothermal Amplification (LAMP) assays for detection of these pathogens.Methods and MaterialsThe LAMP assays, performed in a Genie III real-time fluorometer, targeted a 263 bp region of theC. psittaci-specific Cps_0607 gene or a 209 bp region of aC. pecorum-specific conserved gene CpecG_0573, and were evaluated using a range of samples previously screened using species-specific quantitative PCRs (qPCRs). Species-specificity forC. psittaciandC. pecorumLAMP targets was tested against DNA samples from related chlamydial species and a range of other bacteria. In order to evaluate pathogen detection in clinical samples,C. psittaciLAMP was evaluated using a total of 26 DNA extracts from clinical samples from equine and avian hosts, while forC. pecorumLAMP, we tested a total of 63 DNA extracts from clinical samples from koala, sheep and cattle hosts. A subset of 36C. pecorumsamples was also tested in a thermal cycler (instead of a real-time fluorometer) using newly developed LAMP and results were determined as an end point detection. We also evaluated rapid swab processing (without DNA extraction) to assess the robustness of these assays.ResultsBoth LAMP assays were demonstrated to species-specific, highly reproducible and to be able to detect as little as 10 genome copy number/reaction, with a mean amplification time of 14 and 24 min forC. psittaciandC. pecorum, respectively. When testing clinical samples, the overall congruence between the newly developed LAMP assays and qPCR was 92.3% forC. psittaci(91.7% sensitivity and 92.9% specificity); and 84.1% forC. pecorum(90.6% sensitivity and 77.4% specificity). For a subset of 36C. pecorumsamples tested in a thermal cycler using newly developed LAMP, we observed 34/36 (94.4%) samples result being congruent between LAMP performed in fluorometer and in thermal cycler. Rapid swab processing method evaluated in this study also allows for chlamydial DNA detection using LAMP.DiscussionIn this study, we describe the development of novel, rapid and robustC. psittaci-specific andC. pecorum-specific LAMP assays that are able to detect these bacteria in clinical samples in either the laboratory or POC settings. With further development and a focus on the preparation of these assays at the POC, it is anticipated that both tests may fill an important niche in the repertoire of ancillary diagnostic tools available to clinicians.

scholarly journals A miniaturized optoelectronic biosensor for real-time point-of-care total protein analysis.

MethodsX ◽  
2021 ◽  
pp. 101414
Author(s):  
Ophir Vermesh ◽  
Fariah Mahzabeen ◽  
Jelena Levi ◽  
Marilyn Tan ◽  
Israt S. Alam ◽  
...  
Keyword(s):  
Real Time ◽  
Total Protein ◽  
Point Of Care ◽  
Protein Analysis ◽  
Time Point

scholarly journals Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Yang Zhang ◽  
Chunyang Dai ◽  
Huiyan Wang ◽  
Yong Gao ◽  
Tuantuan Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Highly Sensitive ◽  
One Step ◽  
Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

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