scholarly journals A fully integrated paperfluidic molecular diagnostic chip for the extraction, amplification, and detection of nucleic acids from clinical samples

Lab on a Chip ◽  
2016 ◽  
Vol 16 (4) ◽  
pp. 753-763 ◽  
Author(s):  
Natalia M. Rodriguez ◽  
Winnie S. Wong ◽  
Lena Liu ◽  
Rajan Dewar ◽  
Catherine M. Klapperich
Keyword(s):  
Nucleic Acids ◽  
Point Of Care ◽  
Low Cost ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Fully Integrated

We present a low-cost, disposable, and fully-integrated paperfluidic molecular diagnostic chip for sample-to-result functionality at the point-of-care.

A point-of-care diagnostic for differentiating Ebola from endemic febrile diseases

2018 ◽  
Vol 10 (471) ◽  
pp. eaat0944 ◽  
Author(s):  
David Sebba ◽  
Alexander G. Lastovich ◽  
Melody Kuroda ◽  
Eric Fallows ◽  
Joshua Johnson ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Clinical Symptoms ◽  
Point Of Care ◽  
Hemorrhagic Fever ◽  
Diagnostic Methods ◽  
Outbreak Detection ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Live Virus ◽  
And Control

Hemorrhagic fever outbreaks such as Ebola are difficult to detect and control because of the lack of low-cost, easily deployable diagnostics and because initial clinical symptoms mimic other endemic diseases such as malaria. Current molecular diagnostic methods such as polymerase chain reaction require trained personnel and laboratory infrastructure, hindering diagnostics at the point of need. Although rapid tests such as lateral flow can be broadly deployed, they are typically not well-suited for differentiating among multiple diseases presenting with similar symptoms. Early detection and control of Ebola outbreaks require simple, easy-to-use assays that can detect and differentiate infection with Ebola virus from other more common febrile diseases. Here, we developed and tested an immunoassay technology that uses surface-enhanced Raman scattering (SERS) tags to simultaneously detect antigens from Ebola, Lassa, and malaria within a single blood sample. Results are provided in <30 min for individual or batched samples. Using 190 clinical samples collected from the 2014 West African Ebola outbreak, along with 163 malaria positives and 233 negative controls, we demonstrated Ebola detection with 90.0% sensitivity and 97.9% specificity and malaria detection with 100.0% sensitivity and 99.6% specificity. These results, along with corresponding live virus and nonhuman primate testing of an Ebola, Lassa, and malaria 3-plex assay, indicate the potential of the SERS technology as an important tool for outbreak detection and clinical triage in low-resource settings.

scholarly journals Electrochemical DNA Detection Methods to Measure Circulating Tumour DNA for Enhanced Diagnosis and Monitoring of Cancer

Proceedings ◽  
2020 ◽  
Vol 60 (1) ◽  
pp. 15
Author(s):  
Bukola Attoye ◽  
Matthew Baker ◽  
Chantevy Pou ◽  
Fiona Thomson ◽  
Damion K. Corrigan
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Detection Methods ◽  
Clinical Samples ◽  
Label Free ◽  
Ambient Conditions ◽  
Liquid Biopsies ◽  
Current Time ◽  
Electrochemical Approach ◽  
Label Free Detection

Liquid biopsies are becoming increasingly important as a potential replacement for existing biopsy procedures which can be invasive, painful and compromised by tumour heterogeneity. This paper reports a simple electrochemical approach tailored towards point-of-care cancer detection and treatment monitoring from biofluids using a label-free detection strategy. The mutations under test were the KRAS G12D and G13D mutations, which are both important in the development and progression of many human cancers and which have a presence that correlates with poor outcomes. These common circulating tumour markers were investigated in clinical samples and amplified by standard and specialist PCR methodologies for subsequent electrochemical detection. Following pre-treatment of the sensor to present a clean surface, DNA probes developed specifically for detection of the KRAS G12D and G13D mutations were immobilized onto low-cost carbon electrodes using diazonium chemistry and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide coupling. Following the functionalisation of the sensor, it was possible to sensitively and specifically detect a mutant KRAS G13D PCR product against a background of wild-type KRAS DNA from the representative cancer sample. Our findings give rise to the basis of a simple and very low-cost system for measuring ctDNA biomarkers in patient samples. The current time to result of the system was 3.5 h with considerable scope for optimisation, and it already compares favourably to the UK National Health Service biopsy service where patients can wait weeks for their result. This paper reports the technical developments we made in the production of consistent carbon surfaces for functionalisation, assay performance data for KRAS G13D and detection of PCR amplicons under ambient conditions.

scholarly journals System Architecture for IIoT-Based POC Molecular Diagnostic Device

2021 ◽  
Vol 6 (1) ◽  
pp. 60
Author(s):  
Byeong-Heon Kil ◽  
Ji-Seong Park ◽  
Chan-Young Park ◽  
Yu-Seop Kim ◽  
Jong-Dae Kim
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Diagnostic System ◽  
Pcr Amplification ◽  
Dna Amplification ◽  
System Structure ◽  
World Health ◽  
Molecular Diagnostic ◽  
Target System ◽  
Open Platform

In this paper, we investigate an efficient structure for a point-of-care (POC) molecular diagnostic system based on the industrial Internet of things (IIoT). The target system can perform automated molecular diagnosis including DNA extraction, PCR amplification, and fluorescence detection. Samples and reagents are placed in a multi-room cartridge and loaded into the system. A rotating motor and a syringe motor control the cartridge to extract DNA from the sample. The extracted DNA is transferred to a polymerase chain reaction (PCR) chamber for DNA amplification and detection. The proposed system provides multiplexing of up to four colors. For POC molecular diagnostics, the World Health Organization demands features such as low volume, low cost, fast results, and a user-friendly interface. In this paper, we propose a system structure that can satisfy these requirements by using a PCR chip and open platform. A distributed structure is adopted for the convenience of maintenance, and a web-based GUI is adopted for the user’s convenience. We also investigated communication problems that may occur between system components. Using the proposed structure, the user can conveniently control from standard computing devices including a smartphone.

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 Application of the amplification-free SERS-based CRISPR/Cas12a platform in the identification of SARS-CoV-2 from clinical samples

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Jiajie Liang ◽  
Peijun Teng ◽  
Wei Xiao ◽  
Guanbo He ◽  
Qifang Song ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Point Of Care ◽  
Clinical Samples ◽  
Nasopharyngeal Swab ◽  
Enhancement Effect ◽  
Diagnostic Assays ◽  
Point Of Care Test ◽  
Raman Enhancement ◽  
Potential Strategy ◽  
Refractory Diseases

AbstractThe control of contagious or refractory diseases requires early, rapid diagnostic assays that are simple, fast, and easy-to-use. Here, easy-to-implement CRISPR/Cas12a-based diagnostic platform through Raman transducer generated by Raman enhancement effect, term as SERS-CRISPR (S-CRISPR), are described. The S-CRISPR uses high-activity noble metallic nanoscopic materials to increase the sensitivity in the detection of nucleic acids, without amplification. This amplification-free platform, which can be performed within 30–40 min of incubation time, is then used for detection of SARS-CoV-2 derived nucleic acids in RNA extracts obtained from nasopharyngeal swab specimens (n  =  112). Compared with the quantitative reverse transcription polymerase chain reaction (RT-qPCR), the sensitivity and specificity of S-CRISPR reaches 87.50% and 100%, respectively. In general, the S-CRISPR can rapidly identify the RNA of SARS-CoV-2 RNA without amplification and is a potential strategy for nucleic acid point of care test (POCT).

scholarly journals Highly performing point-of-care molecular testing for SARS-CoV-2 with RNA extraction and isothermal amplification.

2020 ◽  
Author(s):  
Pierre Garneret ◽  
Etienne Coz ◽  
Elian Martin ◽  
Jean-Claude Manuguerra ◽  
Elodie Brient-Litzler ◽  
...  
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Rna Extraction ◽  
Dna Amplification ◽  
Developed Countries ◽  
Temperature Cycling ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Clinical Samples ◽  
Molecular Testing

In order to respond to the urgent request of massive testing, developed countries perform nucleic acid amplification tests (NAAT) of SARS-CoV-2 in centralized laboratories. Real-time RT - PCR (Reverse transcription - Polymerase Chain Reaction) is used to amplify the viral RNA and enable its detection. Although PCR is 37 years old, it is still considered, without dispute, as the gold standard. PCR is an efficient process, but the complex engineering required for automated RNA extraction and temperature cycling makes it incompatible for use in point of care settings. In the present work, by harnessing progress made in the past two decades in DNA amplification, microfluidics and membrane technologies, we succeeded to create a portable test, in which SARS-CoV-2 RNA is extracted, amplified isothermally by RT - LAMP (Loop-mediated Isothermal Amplification), and detected using intercalating dyes or highly fluorescent probes. Depending on the viral load, the detection takes between twenty minutes and one hour. Using pools of naso-pharyngal clinical samples, we estimated a sensitivity comparable to RT-qPCR (up to a Cycle threshold of 39, equivalent to <0.1 TCID50 per mL) and a 100% specificity, for other human coronaviruses and eight respiratory viruses currently circulating in Europe. We designed and fabricated an easy-to-use portable device called COVIDISC to carry out the test at the point of care. The low cost of the materials along with the absence of complex equipment paves the way towards a large dissemination of this device. The perspective of a reliable SARS-CoV-2 point of care detection, highly performing, that would deliver on-site results in less than one hour opens up a new efficient approach to manage the pandemics.

scholarly journals Detection of ESKAPE Bacterial Pathogens at the Point of Care Using Isothermal DNA-Based Assays in a Portable Degas-Actuated Microfluidic Diagnostic Assay Platform

2016 ◽  
Vol 83 (4) ◽  
Author(s):  
Lars D. Renner ◽  
Jindong Zan ◽  
Linda I. Hu ◽  
Manuel Martinez ◽  
Pedro J. Resto ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Point Of Care ◽  
Low Cost ◽  
Bacterial Pathogens ◽  
Diagnostic System ◽  
Molecular Diagnostic ◽  
Technology Gap ◽  
Content Type ◽  
Antimicrobial Drug Resistance ◽  
Industrial Processing

ABSTRACT An estimated 1.5 billion microbial infections occur globally each year and result in ∼4.6 million deaths. A technology gap associated with commercially available diagnostic tests in remote and underdeveloped regions prevents timely pathogen identification for effective antibiotic chemotherapies for infected patients. The result is a trial-and-error approach that is limited in effectiveness, increases risk for patients while contributing to antimicrobial drug resistance, and reduces the lifetime of antibiotics. This paper addresses this important diagnostic technology gap by describing a low-cost, portable, rapid, and easy-to-use microfluidic cartridge-based system for detecting the ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) bacterial pathogens that are most commonly associated with antibiotic resistance. The point-of-care molecular diagnostic system consists of a vacuum-degassed microfluidic cartridge preloaded with lyophilized recombinase polymerase amplification (RPA) assays and a small portable battery-powered electronic incubator/reader. The isothermal RPA assays detect the targeted ESKAPE pathogens with high sensitivity (e.g., a limit of detection of ∼10 nucleic acid molecules) that is comparable to that of current PCR-based assays, and they offer advantages in power consumption, engineering, and robustness, which are three critical elements required for the point-of-care setting. IMPORTANCE This paper describes a portable system for rapidly identifying bacteria in resource-limited environments; we highlight the capabilities of the technology by detecting different pathogens within the ESKAPE collection, which cause nosocomial infections. The system is designed around isothermal DNA-based assays housed within an autonomous plastic cartridge that are designed with the end user in mind, who may have limited technological training. Displaying excellent sensitivity and specificity, the assay systems that we demonstrate may enable future diagnoses of bacterial infection to guide the development of effective chemotherapies and may have a role in areas beyond health where rapid detection is valuable, including in industrial processing and manufacturing, food security, agriculture, and water quality testing.

scholarly journals Evaluation of the Point of Care Molecular Diagnostic Genedrive HCV ID Kit for the detection of HCV RNA in clinical samples

2020 ◽  
pp. 1-23
Author(s):  
Abhishek Padhi ◽  
Ekta Gupta ◽  
Gaurav Singh ◽  
Reshu Agarwal ◽  
Manoj Kumar Sharma ◽  
...  
Keyword(s):  
Point Of Care ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Hcv Rna

scholarly journals A Single-Bead-Based, Fully Integrated Microfluidic System for High-Throughput CD4+T Lymphocyte Enumeration

2017 ◽  
Vol 23 (2) ◽  
pp. 134-143 ◽  
Author(s):  
Xianbo Qiu ◽  
Junhui Zhang ◽  
Shisong Gong ◽  
Dong Wang ◽  
Shan Qiao ◽  
...  
Keyword(s):  
T Lymphocyte ◽  
High Throughput ◽  
Point Of Care ◽  
Low Cost ◽  
Ccd Camera ◽  
Microfluidic System ◽  
Magnetic Control ◽  
Magnetic Actuation ◽  
Fully Integrated ◽  
Single Bead

A single-bead-based, fully integrated microfluidic system has been developed for high-throughput CD4+T lymphocyte enumeration at point-of-care testing. Instead of directly counting CD4+T lymphocytes, CD4+T lymphocyte enumeration is achieved by quantitatively detecting CD4 antigen from the lysed blood sample with a functionalized polycarbonate single bead based on chemiluminescence. To implement the sandwiched chemiluminescence immunoassay with reduced nonspecific binding, a streamlined microfluidic chip with multiple reaction chambers is developed to allow each reaction step to be completed in an independent chamber where reagent is pre-stored. With simple magnetic control, the single bead with an embedded ferrous core can be consecutively transported between each of two adjacent chambers for different reactions. Meanwhile, enhanced mixing can be achieved by moving the single bead back and forth inside one chamber with magnetic actuation. High-throughput detection can be performed when a linear actuation stage is adopted to introduce synchronous magnetic control to multiple single beads in parallel microfluidic chips. A sensitive charge-coupled device (CCD) camera is adopted for high-throughput chemiluminescence detection from multiple single beads. Experimental results show that with the fully integrated microfluidic system, easy-to-operate, accurate, low-cost, immediate, and high-throughput CD4+T lymphocyte enumeration can be successfully achieved at resource-poor settings.

scholarly journals Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) System for Rapid Viral DNA Sensing

2020 ◽  
Author(s):  
Kenneth N. Hass ◽  
Mengdi Bao ◽  
Qian He ◽  
Myeongkee Park ◽  
Peiwu Qin ◽  
...  
Keyword(s):  
Point Of Care ◽  
Molecular Diagnostic ◽  
Nucleic Acid Detection ◽  
Viral Dna ◽  
Fully Integrated ◽  
Impact System ◽  
Target Dna ◽  
Reporter Probe ◽  
One Step ◽  
Labeled Probe

ABSTRACTA fully Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) system is developed for viral DNA detection. This powerful system is patterned with high-aspect ratio micropillars to enhance reporter probe binding. After surface modification and probe immobilization, CRISPR Cas12a/crRNA complex is injected into the fully enclosed system. With the presence of double-stranded DNA target, the CRISPR enzyme is activated and non-specifically cleaves the ssDNA reporters initially immobilized on the micropillars. This collateral cleavage releases fluorescence dyes into the assay, and the intensity is linearly proportional to the target DNA concentration ranging from 0.1 to 10 nM. Importantly, this system does not rely on traditional dye-quencher labeled probe thus eliminating the fluorescence background presented in the assay. Furthermore, our one-step detection protocol is performed at isothermal conditions (37°C) without using complicated and time-consuming off-chip probe hybridization and denaturation. This miniaturized and fully packed IMPACT chip demonstrates rapid, sensitive, and simple nucleic acid detection and is an ideal candidate for the next generation molecular diagnostic platform for point-of-care (POC) applications, responding to emerging and deadly pathogen outbreaks.

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