scholarly journals Nitrocellulose-bound achromopeptidase for point-of-care nucleic acid tests

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Georgios Chondrogiannis ◽  
Shirin Khaliliazar ◽  
Anna Toldrà ◽  
Pedro Réu ◽  
Mahiar M. Hamedi
Keyword(s):  
Nucleic Acid ◽  
Sample Preparation ◽  
Point Of Care ◽  
Dna Amplification ◽  
Recombinase Polymerase Amplification ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Enzymatic Function ◽  
Modern Biotechnology ◽  
Nucleic Acid Tests

AbstractEnzymes are the cornerstone of modern biotechnology. Achromopeptidase (ACP) is a well-known enzyme that hydrolyzes a number of proteins, notably proteins on the surface of Gram-positive bacteria. It is therefore used for sample preparation in nucleic acid tests. However, ACP inhibits DNA amplification which makes its integration difficult. Heat is commonly used to inactivate ACP, but it can be challenging to integrate heating into point-of-care devices. Here, we use recombinase polymerase amplification (RPA) together with ACP, and show that when ACP is immobilized on nitrocellulose paper, it retains its enzymatic function and can easily and rapidly be activated using agitation. The nitrocellulose-bound ACP does, however, not leak into the solution, preventing the need for deactivation through heat or by other means. Nitrocellulose-bound ACP thus opens new possibilities for paper-based Point-of-Care (POC) devices.

scholarly journals Label-Free Electrochemical Sensor for Rapid Bacterial Pathogen Detection Using Vancomycin-Modified Highly Branched Polymers

Sensors ◽  
2021 ◽  
Vol 21 (5) ◽  
pp. 1872
Author(s):  
Holger Schulze ◽  
Harry Wilson ◽  
Ines Cara ◽  
Steven Carter ◽  
Edward N. Dyson ◽  
...  
Keyword(s):  
Pathogen Detection ◽  
Electrochemical Impedance ◽  
Point Of Care ◽  
Branched Polymers ◽  
Label Free ◽  
Conformation Change ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Staphylococcus Carnosus ◽  
Label Free Detection

Rapid point of care tests for bacterial infection diagnosis are of great importance to reduce the misuse of antibiotics and burden of antimicrobial resistance. Here, we have successfully combined a new class of non-biological binder molecules with electrochemical impedance spectroscopy (EIS)-based sensor detection for direct, label-free detection of Gram-positive bacteria making use of the specific coil-to-globule conformation change of the vancomycin-modified highly branched polymers immobilized on the surface of gold screen-printed electrodes upon binding to Gram-positive bacteria. Staphylococcus carnosus was detected after just 20 min incubation of the sample solution with the polymer-functionalized electrodes. The polymer conformation change was quantified with two simple 1 min EIS tests before and after incubation with the sample. Tests revealed a concentration dependent signal change within an OD600 range of Staphylococcus carnosus from 0.002 to 0.1 and a clear discrimination between Gram-positive Staphylococcus carnosus and Gram-negative Escherichia coli bacteria. This exhibits a clear advancement in terms of simplified test complexity compared to existing bacteria detection tests. In addition, the polymer-functionalized electrodes showed good storage and operational stability.

Achieving room temperature DNA amplification by dialling in destabilization

2015 ◽  
Vol 51 (44) ◽  
pp. 9101-9104 ◽  
Author(s):  
B. Safeenaz Alladin-Mustan ◽  
Catherine J. Mitran ◽  
Julianne M. Gibbs-Davis
Keyword(s):  
Nucleic Acid ◽  
Room Temperature ◽  
Point Of Care ◽  
Dna Amplification ◽  
Heating Element ◽  
Nucleic Acid Sequences

The ability to amplify nucleic acid sequences at room temperature without the need for any heating element has been achieved, which has promise in bio-diagnostics employed at the point of care.

Different Methods for Molecular and Rapid Detection of Human Novel Coronavirus

2021 ◽  
Vol 27 ◽  
Author(s):  
Mohamad Hesam Shahrajabian ◽  
Wenli Sun ◽  
Qi Cheng
Keyword(s):  
Nucleic Acid ◽  
Rapid Detection ◽  
Pathogenic Bacteria ◽  
Scientific Information ◽  
Meat Products ◽  
Lamp Assay ◽  
Dna Amplification ◽  
Digital Pcr ◽  
Recombinase Polymerase Amplification ◽  
Nucleic Acid Amplification

Introduction: While PCR has been recognized as one of the appropriate ways to diagnosis of infectious diseases, Loop-mediated isothermal amplification (LAMP), which is a nucleic acid amplification method, can be considered as an alternative to PCR, and it is faster, cost-effective, and easier to perform than nested PCR. Patients and Methods: Keywords were searched in PubMed/MEDLINE, Scopus and Institute for Scientific Information Web of Science, as well as the search engine of Google Scholar. Keywords were PCR, LAMP, RAA, RPA, Virus and COVID-19. Results: LAMP technology has been extensively applied for the detection of human pathogenic bacteria, crop pests, pathogenic organisms and components in meat products. A new isotheral method, Recombinase polymerase amplification (RPA), can amplify the DNA as well as RPA. RPA has benefited from isothermal PCR and both simplicity and rapid amplification. Recombinase aided amplification (RAA) assay has been favorably used in the detection of bacterial and viral pathogens and solved the technical difficulties posed by DNA amplification methods because it does not need thermal denaturation of the template and employs at a low and constant temperature. Conclusions: Reverse transcription polymerase chain reaction, digital PCR, LAMP, nicking endonuclease amplification reaction, recombinase polymerase amplification, and clustered regularly interspaced short palindromic repeats are different nucleic acid amplification tests of COVID-19. LAMP methods can be more specific than qPCR and immunoassays. The LAMP assay can be applied for rapid detection of SARS-CoV, MERS-CoV, SARS-CoV-2, and influenza, because LAMP is a highly sensitive and specific DNA/RNA amplification technique.

Current State of Commercial Point-of-Care Nucleic Acid Tests for Infectious Diseases

The Analyst ◽  
2021 ◽  
Author(s):  
Jane Zhang ◽  
Andrew T. Bender ◽  
David S. Boyle ◽  
Paul K Drain ◽  
Jonathan D. Posner
Keyword(s):  
Nucleic Acid ◽  
Infectious Diseases ◽  
Point Of Care ◽  
Current State ◽  
Nucleic Acid Tests

The COVID-19 pandemic has put the spotlight on the urgent need for integrated nucleic acid tests (NATs) for infectious diseases, especially those that can be used near patient (“point-of-care”, POC),...

scholarly journals Towards a “Sample-In, Answer-Out” Point-of-Care Platform for Nucleic Acid Extraction and Amplification: Using an HPV E6/E7 mRNA Model System

2012 ◽  
Vol 2012 ◽  
pp. 1-12 ◽  
Author(s):  
Anja Gulliksen ◽  
Helen Keegan ◽  
Cara Martin ◽  
John O'Leary ◽  
Lars A. Solli ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Sample Preparation ◽  
Clinical Sample ◽  
Point Of Care ◽  
Fluorescent Detection ◽  
Clinical Samples ◽  
Microfluidic Channels ◽  
Acid Extraction ◽  
Nucleic Acid Extraction ◽  
Hpv E6

The paper presents the development of a “proof-of-principle” hands-free and self-contained diagnostic platform for detection of human papillomavirus (HPV) E6/E7 mRNA in clinical specimens. The automated platform performs chip-based sample preconcentration, nucleic acid extraction, amplification, and real-time fluorescent detection with minimal user interfacing. It consists of two modular prototypes, one for sample preparation and one for amplification and detection; however, a common interface is available to facilitate later integration into one single module. Nucleic acid extracts (n=28) from cervical cytology specimens extracted on the sample preparation chip were tested using the PreTect HPV-Proofer and achieved an overall detection rate for HPV across all dilutions of 50%–85.7%. A subset of 6 clinical samples extracted on the sample preparation chip module was chosen for complete validation on the NASBA chip module. For 4 of the samples, a 100% amplification for HPV 16 or 33 was obtained at the 1 : 10 dilution for microfluidic channels that filled correctly. The modules of a “sample-in, answer-out” diagnostic platform have been demonstrated from clinical sample input through sample preparation, amplification and final detection.

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