Cargando…

Maximising the Use of Scarce qPCR Master Mixes

The COVID-19 pandemic resulted in a universal, immediate, and vast demand for comprehensive molecular diagnostic testing, especially real-time quantitative (qPCR)-based methods. This rapidly triggered a global shortage of testing capacity, equipment, and reagents. Even today, supply times for chemic...

Descripción completa

Detalles Bibliográficos
Autores principales: Bustin, Stephen, Bustin, Claire, Kirvell, Sara, Nolan, Tania, Mueller, Reinhold, Shipley, Gregory
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9368830/
https://www.ncbi.nlm.nih.gov/pubmed/35955620
http://dx.doi.org/10.3390/ijms23158486
_version_ 1784766263726178304
author Bustin, Stephen
Bustin, Claire
Kirvell, Sara
Nolan, Tania
Mueller, Reinhold
Shipley, Gregory
author_facet Bustin, Stephen
Bustin, Claire
Kirvell, Sara
Nolan, Tania
Mueller, Reinhold
Shipley, Gregory
author_sort Bustin, Stephen
collection PubMed
description The COVID-19 pandemic resulted in a universal, immediate, and vast demand for comprehensive molecular diagnostic testing, especially real-time quantitative (qPCR)-based methods. This rapidly triggered a global shortage of testing capacity, equipment, and reagents. Even today, supply times for chemicals from date of order to delivery are often much longer than pre-pandemic. Furthermore, many companies have ratcheted up the price for minimum volumes of reaction master mixes essential for qPCR assays, causing additional problems for academic laboratories often operating on a shoestring. We have validated two strategies that stretch reagent supplies and, whilst particularly applicable in case of scarcity, can readily be incorporated into standard qPCR protocols, with appropriate validation. The first strategy demonstrates equivalent performance of a selection of “past expiry date” and newly purchased master mixes. This approach is valid for both standard and fast qPCR protocols. The second validates the use of these master mixes at less than 1x final concentration without loss of qPCR efficiency or sensitivity.
format Online
Article
Text
id pubmed-9368830
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-93688302022-08-12 Maximising the Use of Scarce qPCR Master Mixes Bustin, Stephen Bustin, Claire Kirvell, Sara Nolan, Tania Mueller, Reinhold Shipley, Gregory Int J Mol Sci Technical Note The COVID-19 pandemic resulted in a universal, immediate, and vast demand for comprehensive molecular diagnostic testing, especially real-time quantitative (qPCR)-based methods. This rapidly triggered a global shortage of testing capacity, equipment, and reagents. Even today, supply times for chemicals from date of order to delivery are often much longer than pre-pandemic. Furthermore, many companies have ratcheted up the price for minimum volumes of reaction master mixes essential for qPCR assays, causing additional problems for academic laboratories often operating on a shoestring. We have validated two strategies that stretch reagent supplies and, whilst particularly applicable in case of scarcity, can readily be incorporated into standard qPCR protocols, with appropriate validation. The first strategy demonstrates equivalent performance of a selection of “past expiry date” and newly purchased master mixes. This approach is valid for both standard and fast qPCR protocols. The second validates the use of these master mixes at less than 1x final concentration without loss of qPCR efficiency or sensitivity. MDPI 2022-07-30 /pmc/articles/PMC9368830/ /pubmed/35955620 http://dx.doi.org/10.3390/ijms23158486 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Technical Note
Bustin, Stephen
Bustin, Claire
Kirvell, Sara
Nolan, Tania
Mueller, Reinhold
Shipley, Gregory
Maximising the Use of Scarce qPCR Master Mixes
title Maximising the Use of Scarce qPCR Master Mixes
title_full Maximising the Use of Scarce qPCR Master Mixes
title_fullStr Maximising the Use of Scarce qPCR Master Mixes
title_full_unstemmed Maximising the Use of Scarce qPCR Master Mixes
title_short Maximising the Use of Scarce qPCR Master Mixes
title_sort maximising the use of scarce qpcr master mixes
topic Technical Note
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9368830/
https://www.ncbi.nlm.nih.gov/pubmed/35955620
http://dx.doi.org/10.3390/ijms23158486
work_keys_str_mv AT bustinstephen maximisingtheuseofscarceqpcrmastermixes
AT bustinclaire maximisingtheuseofscarceqpcrmastermixes
AT kirvellsara maximisingtheuseofscarceqpcrmastermixes
AT nolantania maximisingtheuseofscarceqpcrmastermixes
AT muellerreinhold maximisingtheuseofscarceqpcrmastermixes
AT shipleygregory maximisingtheuseofscarceqpcrmastermixes