Role model
Efficiency correction is required for accurate quantitative PCR analysis and reporting./Ruyter, Jan M.; Barnewall, Rebecca J.; Marsh, Ian B. et al.
U:Clinical chemistry, Volume 67, No. 6, 06.01.2021, p. 829-842.
Research:Contribution to the magazine›Article›Academic›peer review
Harvard
Ruyter, JM, Barnewall, RJ, Marsh, IB, Szentirmay, AN, Quinn, JC, likes, R, service, QD2021, 'Efficiency correction is required for accurate quantitative PCR analysis and reporting',Clinical chemistry, sv. 67, br. 6, str. 829-842.https://doi.org/10.1093/clinchem/hvab052
WHAT
Ruijter, J.M., Barnewall, R.J., Marsh, Ι.Β., Szentirmay, A.N., Quinn, J.C., likes, R., Naklonost, Q.D.(2021).Efficiency correction is required for accurate quantitative PCR analysis and reporting.Clinical chemistry,67(6), 829-842.https://doi.org/10.1093/clinchem/hvab052
Vancouver
Ruyter JM, Barnewall RJ, Marsh IB, Szentirmay AN, Quinn JC, van Houd Ret al.Efficiency correction is required for accurate quantitative PCR analysis and reporting.Clinical chemistry. 2021 Jun 1;67(6):829-842. doi: 10.1093/clinchem/hvab052
Author
Ruyter, Jan M.; Barnewall, Rebecca J.; Marsh, Ian B. et al. /Efficiency correction is required for accurate quantitative PCR analysis and reporting. U:Clinical chemistry. 2021; volume 67, no. 6. p. 829-842.
BibTeX
@članak{d64e0cd1cdc241328dc55d76dc11b9ee,
title = "Efficiency correction required for accurate quantitative PCR analysis and reporting",
abstract = "BACKGROUND: Quantitative PCR (qPCR) aims to measure the concentration of DNA or RNA in diagnostic and biological samples based on quantitative cycle time (Cq) values observed in amplification curves. The results of qPCR experiments are routinely calculated as if all assays were 100% efficient or reported only as Cq, ΔCq, or ΔΔCq values. CONTENT: When a reaction shows specific amplification, it should be considered positive, regardless of the observed Cq. Since Cq is highly dependent on amplification efficiency, which can vary between targets and samples, accurate calculation of the target amount and relative gene expression requires consideration of actual amplification efficiency in assays and reports PCR efficiency is often derived from standard curves, but this approach is subject to dilution errors and interferes with these factors affecting the accurate quantification of clinical and biological samples used in diagnostic applications and collected under harsh conditions PCR yields determined from individual amplification curves avoid these confounding factors. To obtain unbiased results with performance correction, we recommend absolute quantification with a single undiluted calibrator of known target concentration and efficiency values derived from amplification curves of calibrators and unknown samples. ABSTRACT: For meaningful diagnosis or biological interpretation, the reported results of qPCR experiments should be corrected for efficiency. To avoid ambiguity, the Minimum Information Guidelines for Publications of Quantitative Real-Time PCR Experiments (MIQE) checklist should be expanded to require the methods used (1) to determine PCR efficiency and (2) to calculate reported target amounts and relative gene expression values.",
keywords = "MIQE, single-point calibration, PCR, absolute quantification, diagnostics, eDNA, efficiency correction, point-of-care, quantitative PCR",
συγγραφέας = "Ruijter, {Jan M.} i Barnewall, {Rebecca J.} i Marsh, {Ian B.} i Szentirmay, {Andrew N.} i Quinn, {Jane C.} i {van Houdt}, Robin i Gunst, {Quinn D.} i {van den Hoff}, {Maurice J.B.}",
note = "Copyright Publisher: {\textcopyright} American Society for Clinical Chemistry 2021.",
year = "2021",
month = June,
and = "1",
doi = "10.1093/clinchem/hvab052",
language = "english",
volume = "67",
pages = "829--842",
journal = "Clinical Chemistry",
issn = "0009-9147",
εκδοτες = "American Society for Clinical Chemistry Inc.",
number = "6",
}
MINISTRY OF
TY-DAN
T1 - Efficiency correction required for accurate quantitative PCR analysis and reporting
AU - Ruijter, Jan M.
AU - Barnewall, Rebecca J.
AU - Marsh, Ian B.
AU - Szentirmay, Andrew N.
AU - Quinn, Jane S.
AU - I like it, Robin
AU - Gunst, Quinn D.
AU - van den Hoff, Maurice J.B.
N1 - Publisher Copyright: © American Society for Clinical Chemistry 2021.
PY - 1.6.2021
Y1 - 1.6.2021
N2 - BACKGROUND: Quantitative PCR (qPCR) aims to measure the concentration of DNA or RNA in diagnostic and biological samples based on the quantification cycle (Cq) value observed in the amplification curves. The results of qPCR experiments are routinely calculated as if all assays were 100% efficient or reported only as Cq, ΔCq, or ΔΔCq values. CONTENT: When a reaction shows specific amplification, it should be considered positive, regardless of the observed Cq. Because Cq is highly dependent on amplification efficiency, which can vary between targets and samples, accurate calculation of target abundance and relative gene expression requires consideration of actual amplification efficiency in analysis and reporting. PCR efficiency is often derived from standard curves, but this approach is affected by dilution errors and hampered by the properties of standards and diluents. These factors affect the accurate quantification of clinical and biological samples used in diagnostic applications and collected under harsh conditions. PCR yields determined from individual amplification curves avoid these confounding factors. To obtain unbiased results with performance correction, we recommend absolute quantification with a single undiluted calibrator of known target concentration and efficiency values derived from amplification curves of calibrators and unknown samples. ABSTRACT: For meaningful diagnosis or biological interpretation, the reported results of qPCR experiments should be corrected for efficiency. To avoid ambiguity, the Minimum Information Guidelines for Publications of Quantitative Real-Time PCR Experiments (MIQE) checklist should be expanded to require the methods used (1) to determine PCR efficiency and (2) to calculate reported target amounts and relative gene expression values.
AB - BACKGROUND: Quantitative PCR (qPCR) aims to measure the concentration of DNA or RNA in diagnostic and biological samples based on the quantification cycle (Cq) value observed in the amplification curves. The results of qPCR experiments are routinely calculated as if all assays were 100% efficient or reported only as Cq, ΔCq, or ΔΔCq values. CONTENT: When a reaction shows specific amplification, it should be considered positive, regardless of the observed Cq. Because Cq is highly dependent on amplification efficiency, which can vary between targets and samples, accurate calculation of target abundance and relative gene expression requires consideration of actual amplification efficiency in analysis and reporting. PCR efficiency is often derived from standard curves, but this approach is affected by dilution errors and hampered by the properties of standards and diluents. These factors affect the accurate quantification of clinical and biological samples used in diagnostic applications and collected under harsh conditions. PCR yields determined from individual amplification curves avoid these confounding factors. To obtain unbiased results with performance correction, we recommend absolute quantification with a single undiluted calibrator of known target concentration and efficiency values derived from amplification curves of calibrators and unknown samples. ABSTRACT: For meaningful diagnosis or biological interpretation, the reported results of qPCR experiments should be corrected for efficiency. To avoid ambiguity, the Minimum Information Guidelines for Publications of Quantitative Real-Time PCR Experiments (MIQE) checklist should be expanded to require the methods used (1) to determine PCR efficiency and (2) to calculate reported target amounts and relative gene expression values.
KW - MIQE
KW - Single point calibration
KW - PCR
KW - absolute quantification
KW - diagnostics
KW - eDNA
KW - performance correction
KW - point of concern
KW - quantitative PCR
UR - http://www.scopus.com/inward/record.url?scp=85107390313&partnerID=8YFLogxK
U2 - 10.1093/clinchem/hvab052
DO - 10.1093/clinchem/hvab052
M3 - Article
C2 - 33890632
VL - 67 (view, expert).
SP - 829 (view, professional).
EP - 842
JO - Clinical chemistry
JF - Clinical chemistry
SN - 0009-9147
JE - 6
IS -