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Naslov:Droplet volume variability as a critical factor for accuracy of absolute quantification using droplet digital PCR
Avtorji:ID Bogožalec Košir, Alexandra (Avtor)
ID Divieto, Carla (Avtor)
ID Pavšič, Jernej (Avtor)
ID Pavarelli, Stefano (Avtor)
ID Dobnik, David (Avtor)
ID Dreo, Tanja (Avtor)
ID Bellotti, Roberto (Avtor)
ID Sassi, Maria Paola (Avtor)
ID Žel, Jana (Avtor)
Datoteke:.pdf PDF - Predstavitvena datoteka, prenos (565,33 KB)
MD5: BEA796FB4D18CADAD0DF006516470C12
 
URL URL - Izvorni URL, za dostop obiščite https://doi.org/10.1007/s00216-017-0625-y
 
Jezik:Angleški jezik
Tipologija:1.01 - Izvirni znanstveni članek
Organizacija:Logo NIB - Nacionalni inštitut za biologijo
Povzetek:Accurate and precise nucleic-acid quantification is crucial for clinical and diagnostic decisions, as overestimation or underestimation can lead to misguided treatment of a disease or incorrect labelling of the products. Digital PCR is one of the best tools for absolute nucleic-acid copy-number determination. However, digital PCR needs to be well characterised in terms of accuracy and sources of uncertainty. With droplet digital PCR, discrepancies between the droplet volume assigned by the manufacturer and measured by independent laboratories have already been shown in previous studies. In the present study, we report on the results of an inter-laboratory comparison of different methods for droplet volume determination that is based on optical microscopy imaging and is traceable to the International System of Units. This comparison was conducted on the same DNA material, with the examination of the influence of parameters such as droplet generators, supermixes, operators, inter-cartridge and intra-cartridge variability, and droplet measuring protocol. The mean droplet volume was measured using a QX200™ AutoDG™ Droplet Digital™ PCR system and two QX100™ Droplet Digital™ PCR systems. The data show significant volume differences between these two systems, as well as significant differences in volume when different supermixes are used. We also show that both of these droplet generator systems produce droplets with significantly lower droplet volumes (13.1%, 15.9%, respectively) than stated by the manufacturer and previously measured by other laboratories. This indicates that to ensure precise quantification, the droplet volumes should be assessed for each system.
Ključne besede:droplet digital PCR, droplet volume, DNA quantification, optical microscopy imaging
Status publikacije:Objavljeno
Verzija publikacije:Objavljena publikacija
Datum objave:17.09.2017
Leto izida:2017
Št. strani:str. 6689-6697
Številčenje:Vol. 409, iss. 28
PID:20.500.12556/DiRROS-19690 Novo okno
UDK:577.2
ISSN pri članku:1618-2642
DOI:10.1007/s00216-017-0625-y Novo okno
COBISS.SI-ID:4421711 Novo okno
Datum objave v DiRROS:25.07.2024
Število ogledov:515
Število prenosov:335
Metapodatki:XML DC-XML DC-RDF
:
BOGOŽALEC KOŠIR, Alexandra, DIVIETO, Carla, PAVŠIČ, Jernej, PAVARELLI, Stefano, DOBNIK, David, DREO, Tanja, BELLOTTI, Roberto, SASSI, Maria Paola in ŽEL, Jana, 2017, Droplet volume variability as a critical factor for accuracy of absolute quantification using droplet digital PCR. Analytical and bioanalytical chemistry [na spletu]. 2017. Vol. 409, no. 28, p. 6689–6697. [Dostopano 2 april 2025]. DOI 10.1007/s00216-017-0625-y. Pridobljeno s: https://dirros.openscience.si/IzpisGradiva.php?lang=slv&id=19690
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Gradivo je del revije

Naslov:Analytical and bioanalytical chemistry
Skrajšan naslov:Anal. bioanal. chem.
Založnik:Springer
ISSN:1618-2642
COBISS.SI-ID:6966294 Novo okno

Gradivo je financirano iz projekta

Financer:ARIS - Javna agencija za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije
Številka projekta:P4-0165-2015
Naslov:Biotehnologija in sistemska biologija rastlin
Financer:EC - European Commission
Program financ.:European Metrology Research Programme (EMRP)
Številka projekta:JRP SIB54
Naslov:Traceability for biologically relevant molecules and entities
Akronim:Bio-SITrace
Financer:Drugi - Drug financer ali več financerjev
Program financ.:EURAMET and the European Union
Akronim:EMRP
Financer:ARIS - Javna agencija za znanstvenoraziskovalno in inovacijsko dejavnost Republike Slovenije
Številka projekta:1000-15-0105

Licence

Licenca:CC BY 4.0, Creative Commons Priznanje avtorstva 4.0 Mednarodna
Povezava:http://creativecommons.org/licenses/by/4.0/deed.sl
Opis:To je standardna licenca Creative Commons, ki daje uporabnikom največ možnosti za nadaljnjo uporabo dela, pri čemer morajo navesti avtorja.

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