We use cookies to understand how you use our site and to improve your experience. This includes personalizing content and advertising. To learn more, click here. By continuing to use our site, you accept our use of cookies. Cookie Policy.

Features Partner Sites Information LinkXpress
Sign In
Advertise with Us
LGC Clinical Diagnostics


Thermo Fisher Scientific provides analytical instruments, lab equipment, specialty diagnostics, reagents and integrat... read more Featured Products: More products

Download Mobile App

Direct Real-Time PCR Protocol Detects Monkeypox Virus

By LabMedica International staff writers
Posted on 16 Nov 2022
Print article
Image: The Applied Biosystems Quant Studio 6 Pro Real-Time PCR System adds innovative smart features to the real-time PCR (qPCR) workflow, all in a compact footprint (Photo courtesy of Thermo Fisher Scientific)
Image: The Applied Biosystems Quant Studio 6 Pro Real-Time PCR System adds innovative smart features to the real-time PCR (qPCR) workflow, all in a compact footprint (Photo courtesy of Thermo Fisher Scientific)

Monkeypox virus, an encapsulated double-stranded DNA virus and member of the Poxviridae family, is responsible for the recent monkeypox outbreak that has been declared a public health emergency of international concern.

Prompt identification of infected individuals followed by contact tracing is important for stemming the spread of disease. The characteristic rash of monkeypox progresses through multiple stages, beginning with a macular phase, progressing through papular, vesicular, and pustular phases, and ending with a scab phase.

Clinical Pathologists at the Feinberg School of Medicine (Chicago, IL, USA) collected clinical specimens from patients at locations within the Northwestern Medicine health system. Lesions were swabbed with sterile synthetic swabs, and the swabs were submitted to the laboratory dry or in 3 mL of viral transport media (M4 VTM). Dry swabs received by the laboratory were immediately added to 3 mL of M4 VTM. At the start of the monkeypox outbreak, a total of 20 samples identified as positive by the direct assay and 20 samples identified as negative by the direct assay were sequentially chosen for confirmation by indirect method. DNA extraction for the indirect method was performed using the Qiagen manual DNA extraction kit utilizing spin-column–based nucleic acid purification (Qiagen, Germantown MD, USA).

A modified multiplex version of the CDC monkeypox assay was performed for clinical validation purposes. Previously published probe and primers targeting monkeypox were used. After processing, this was followed by real-time PCR on the Quant Studio 6 instrument (Thermo Fisher Scientific, Waltham, MA, USA). Cycling conditions included a 20-second activation step at 95 °C, followed by 40 cycles of 3 seconds at 95 °C and 30 seconds at 60°C.

The investigators generated a standard curve was by diluting plasmid monkeypox control DNA to concentrations ranging from 1 to 1,000,000 copies/mL and determining the corresponding CT value. The assay displayed excellent linearity (R2 = 0.9994). The limit of detection was determined by replicate determinations of CT values (n = 20) of 5, 50, and 1000 copies/mL samples. The mean CT values of 5 copies/mL were determined to be 36 on both the direct and indirect assay, with an SD of 0.75 (range, 34.61 to 37.39). The analytical specificity was determined by running the assay with control materials for 23 different viruses, bacteria, and fungi. No signal within the limit of detection was detected by the assay in any of the control materials. Blood did have an inhibitory effect on the assay, with increasing concentration of blood leading to greater inhibition. Samples with 20% blood had complete inhibition.

The authors concluded that the validation of a direct method monkeypox assay will allow laboratories to lower costs, reduce dependence on the supply chain for nucleic acid extraction kits, and decrease exposure of laboratory scientists to potentially infectious specimens. In addition, it may be suitable for incorporation into automated and high-throughput testing. This direct method will make it easier for laboratories across the world to rapidly develop, validate, and scale testing for monkeypox virus. The study was published in the November 2022 issue of The Journal of Molecular Diagnostics.

Related Links:
Feinberg School of Medicine
Thermo Fisher Scientific

Platinum Supplier
Xylazine Immunoassay Test
Xylazine ELISA
Gold Supplier
Automated, Random Access Chemistry Analyzer
LIDA 300
Gold Supplier
Pipette Controller
Sapphire MaxiPette
Ferritin Test

Print article


Clinical Chemistry

view channel
Image: The new assays are designed to run on the B•R•A•H•M•S KRYPTOR compact PLUS clinical chemistry analyzer (Photo courtesy of Thermo Fisher)

Breakthrough Immunoassays to Aid in Risk Assessment of Preeclampsia

Preeclampsia is a life-threatening blood pressure condition that can arise during pregnancy and the postpartum phase. This severe pregnancy complication is a primary cause of maternal and fetal mortality... Read more


view channel
Image: The latest FDA clearance has finally brought HemoScreen to its full potential as a true POC hematology analyzer (Photo courtesy of PixCell)

True POC Hematology Analyzer with Direct Capillary Sampling Enhances Ease-of-Use and Testing Throughput

An innovative 5-part differential Complete Blood Count (CBC) analyzer with direct capillary sampling capability significantly simplifies blood sampling and minimizes the pre-analytical process.... Read more


view channel
Image: A new test could detect the body’s adaptive immune response to viruses (Photo courtesy of 123RF)

Predictive T-Cell Test Detects Immune Response to Viruses Even Before Antibodies Form

The adaptive immune system is an incredible defense mechanism that allows the human body to identify and mount targeted responses against specific pathogens. T-Cells, a special kind of white blood cell,... Read more


view channel
Image: On-chip pretreatment and rapid AST based directly on positive blood cultures (Photo courtesy of Liu Yang)

Integrated Solution for Rapid AST Directly From Positive Blood Cultures to Combat Bloodstream Infection

The presence of living bacteria in the bloodstream, known as bacteremia, can lead to serious conditions like bloodstream infections (BSIs) and sepsis, which can often be fatal. Quickly prescribing the... Read more


view channel
Image: The Mitotic Counting algorithm was trained on over 100,000 individual mitosis and across seven scanner models (Photo courtesy of Gestalt Diagnostics)

New AI Algorithm for Use on Eight Types of Cancer Trained on Seven Scanner Models

A new artificial intelligence (AI) algorithm for use on one of the broadest trained environments is now available for commercial or research work. Gestalt Diagnostics (Spokane, WA, USA) has released the... Read more


view channel
Image: A new electrochemical device can quickly and inexpensively identify people at greatest risk for osteoporosis (Photo courtesy of ACS Central Science, 2023)

Electrochemical Device Identifies People at Higher Risk for Osteoporosis Using Single Blood Drop

With the global increase in life expectancy, the incidence of age-related conditions like osteoporosis is increasing. Osteoporosis, affecting around 200 million individuals worldwide, has a higher incidence... Read more


view channel
Image: AACC Middle East is a two-day conference that brings the latest in laboratory medicine to the Middle East region (Photo courtesy of ADLM)

AACC Middle East 2023 to Explore Latest Trends in Clinical Pathology and Laboratory Medicine

The AACC Middle East Conference and Exposition will be held by the Association for Diagnostics & Laboratory Medicine (ADLM - formerly AACC, Washington, DC, USA) in partnership with Life Dx (Abu Dhabi,... Read more
Copyright © 2000-2023 Globetech Media. All rights reserved.