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Droplet volume variability as a critical factor for accuracy of absolute quantification using droplet digital PCR
Alexandra Bogožalec Košir, Carla Divieto, Jernej Pavšič, Stefano Pavarelli, David Dobnik, Tanja Dreo, Roberto Bellotti, Maria Paola Sassi, Jana Žel, 2017, original scientific article

Abstract: 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.
Keywords: droplet digital PCR, droplet volume, DNA quantification, optical microscopy imaging
Published in DiRROS: 25.07.2024; Views: 121; Downloads: 78
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Why are the Early Gothic murals in St. Jacob’s Church in Ormož, Slovenia, almost entirely black?
Anabelle Križnar, Katja Kavkler, Sabina Dolenec, 2024, original scientific article

Abstract: In St. Jacob’s parish church in Ormož, Slovenia, mural paintings from around 1350–1370 are partially conserved in the northeastern corner of the main nave. They are almost completely black, indicating a large-scale pigment degradation. They were studied as a part of a larger research project aiming to identify materials applied and their possible degradation. First, they were studied in situ, and next, extracted samples of plaster, pigments, and colour layers were analysed by optical microscopy, Raman spectroscopy, FTIR spectroscopy, SEM-EDS, and XRD. Haematite, green earth, malachite, azurite, and tenorite were identified, showing that azurite and perhaps also malachite degraded to black tenorite, probably due to their fine grinding and their application directly on the fresh plaster. The plaster is made with small and large amounts of aggregate with mostly quartz with some impurities, which makes it fragile. The original appearance of these murals was of bright blue and green colours.
Keywords: mural painting, medieval painting, Gothic art, St. Jacob's Church, Ormož, Slovenia, pigments, chemical structure, colour degradation, optical microscopy, Raman microspectroscopy, FTIR, XRD
Published in DiRROS: 17.04.2024; Views: 250; Downloads: 250
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