Variability and cost implications of three generations of the Roche LightCycler® 480.

2.50
Hdl Handle:
http://hdl.handle.net/10029/621151
Title:
Variability and cost implications of three generations of the Roche LightCycler® 480.
Authors:
Dullaert-de Boer, Maria; Akkerman, Onno W; Vermeer, Marloes; Hess, Dorine L J; Kerstjens, Huib A M; Anthony, Richard M; van der Werf, Tjip S; van Soolingen, Dick; van der Zanden, Adri G M
Abstract:
Real time PCR has become a dominant method for the highly sensitive detection of pathogens in clinical material. Real time PCR can generate a fluorescence signal by using fluorescence labelled probes, allowing us to detect and semi quantify the amount of amplified DNA. Here we test the variability of the detection system and cost implications of three different versions of the LightCycler® 480 (LC480), focusing on the intensity of fluorescence and Cq in monoplex and multiplex rtPCRs. For gastro-intestinal pathogens there was no correlation between the intensity of fluorescence and the Cq value in the different LC480 types. For probes with the dyes FAMTM, HEXTM, Cy5 and Red610 a higher fluorescence intensity was seen in LC480 type II and III compared to LC480 type I. After lowering the probe concentration for the Cy5 dye three-fold (from 0.3μM to 0.1μM) the Cq value remains the same and the intensity of fluorescence decreases. For the LC480 type II and III the difference in fluorescence intensity was much more extreme. The concentration of the different labelled probes can be lowered at least six-fold in LC480 type II and III cyclers while maintaining a fluorescence intensity as high as achieved in the LC480 type I with undiluted probe. In conclusion, the strength of the fluorescence signal of the LightCycler® 480 type III is superior to that of LightCycler® 480 types I and II, allowing the use of lower probe concentrations for all dyes, particularly for the dyes Red610 and Cy5. This results in a two thirds reduction in PCR probe costs. Switching to these newer machines for real-time PCR can reduce dye labelled probe consumption and thus reduce costs significantly.
Citation:
Variability and cost implications of three generations of the Roche LightCycler® 480. 2018, 13 (1):e0190847 PLoS ONE
Journal:
PloS One 2018; 13(1):e190847
Issue Date:
12-Jan-2018
URI:
http://hdl.handle.net/10029/621151
DOI:
10.1371/journal.pone.0190847
PubMed ID:
29329324
Language:
en
ISSN:
1932-6203
Appears in Collections:
Miscellaneous

Full metadata record

DC FieldValue Language
dc.contributor.authorDullaert-de Boer, Mariaen
dc.contributor.authorAkkerman, Onno Wen
dc.contributor.authorVermeer, Marloesen
dc.contributor.authorHess, Dorine L Jen
dc.contributor.authorKerstjens, Huib A Men
dc.contributor.authorAnthony, Richard Men
dc.contributor.authorvan der Werf, Tjip Sen
dc.contributor.authorvan Soolingen, Dicken
dc.contributor.authorvan der Zanden, Adri G Men
dc.date.accessioned2018-01-16T08:55:31Z-
dc.date.available2018-01-16T08:55:31Z-
dc.date.issued2018-01-12-
dc.identifier.citationVariability and cost implications of three generations of the Roche LightCycler® 480. 2018, 13 (1):e0190847 PLoS ONEen
dc.identifier.issn1932-6203-
dc.identifier.pmid29329324-
dc.identifier.doi10.1371/journal.pone.0190847-
dc.identifier.urihttp://hdl.handle.net/10029/621151-
dc.description.abstractReal time PCR has become a dominant method for the highly sensitive detection of pathogens in clinical material. Real time PCR can generate a fluorescence signal by using fluorescence labelled probes, allowing us to detect and semi quantify the amount of amplified DNA. Here we test the variability of the detection system and cost implications of three different versions of the LightCycler® 480 (LC480), focusing on the intensity of fluorescence and Cq in monoplex and multiplex rtPCRs. For gastro-intestinal pathogens there was no correlation between the intensity of fluorescence and the Cq value in the different LC480 types. For probes with the dyes FAMTM, HEXTM, Cy5 and Red610 a higher fluorescence intensity was seen in LC480 type II and III compared to LC480 type I. After lowering the probe concentration for the Cy5 dye three-fold (from 0.3μM to 0.1μM) the Cq value remains the same and the intensity of fluorescence decreases. For the LC480 type II and III the difference in fluorescence intensity was much more extreme. The concentration of the different labelled probes can be lowered at least six-fold in LC480 type II and III cyclers while maintaining a fluorescence intensity as high as achieved in the LC480 type I with undiluted probe. In conclusion, the strength of the fluorescence signal of the LightCycler® 480 type III is superior to that of LightCycler® 480 types I and II, allowing the use of lower probe concentrations for all dyes, particularly for the dyes Red610 and Cy5. This results in a two thirds reduction in PCR probe costs. Switching to these newer machines for real-time PCR can reduce dye labelled probe consumption and thus reduce costs significantly.en
dc.language.isoenen
dc.rightsArchived with thanks to PloS oneen
dc.titleVariability and cost implications of three generations of the Roche LightCycler® 480.en
dc.identifier.journalPloS One 2018; 13(1):e190847en

Related articles on PubMed

All Items in WARP are protected by copyright, with all rights reserved, unless otherwise indicated.