1. In-depth comparison of adeno-associated virus containing fractions after CsCl ultracentrifugation gradient separationMojca Janc, Kaja Zevnik, Ana Dolinar, Tjaša Jakomin, Maja Štalekar, Katarina Bačnik, Denis Kutnjak, Magda Tušek-Žnidarič, Lorena Zentilin, Dmitri G. Fedorov, David Dobnik, 2024, original scientific article Abstract: Recombinant adeno-associated viruses (rAAVs) play a pivotal role in the treatment of genetic diseases. However, current production and purification processes yield AAV-based preparations that often contain unwanted empty, partially filled or damaged viral particles and impurities, including residual host cell DNA and proteins, plasmid DNA, and viral aggregates. To precisely understand the composition of AAV preparations, we systematically compared four different single-stranded AAV (ssAAV) and self-complementary (scAAV) fractions extracted from the CsCl ultracentrifugation gradient using established methods (transduction efficiency, analytical ultracentrifugation (AUC), quantitative and digital droplet PCR (qPCR and ddPCR), transmission electron microscopy (TEM) and enzyme-linked immunosorbent assay (ELISA)) alongside newer techniques (multiplex ddPCR, multi-angle light-scattering coupled to size-exclusion chromatography (SEC-MALS), multi-angle dynamic light scattering (MADLS), and high-throughput sequencing (HTS)). Suboptimal particle separation within the fractions resulted in unexpectedly similar infectivity levels. No single technique could simultaneously provide comprehensive insights in the presence of both bioactive particles and contaminants. Notably, multiplex ddPCR revealed distinct vector genome fragmentation patterns, differing between ssAAV and scAAV. This highlights the urgent need for innovative analytical and production approaches to optimize AAV vector production and enhance therapeutic outcomes.
Keywords: recombinant adeno-associated viruses (rAAVs), CsCl ultracentrifugation gradient, analytical methods, digital droplet PCR (ddPCR), transmission electron microscopy (TEM), analytical ultracentrifugation (AUC), size-exclusion chromatography coupled with multi-angle light scattering (SEC-MALS), Illumina sequencing, virology
Published in DiRROS: 07.08.2024; Views: 92; Downloads: 68
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2. Preverjanje verjetnostne napovedi sanitarnega poseka smreke zaradi podlubnikov v Sloveniji v 2023Nikica Ogris, Maarten De Groot, 2023, other scientific articles Abstract: Preverili smo zanesljivost verjetnostne napovedi sanitarnega poseka smreke zaradi podlubnikov v Sloveniji v 2023. Verjetnostni model za napoved sanitarnega poseka smreke zaradi podlubnikov je potrdil visoko zanesljivost (AUC modela = 0,89, AUC napovedi = 0,84). Ugotovili smo optimalni prag za verjetnost sanitarnega poseka, ki ga bomo lahko uporabili pri naslednjih napovedih za bolj jasno določitev območij, kjer se bodo potencialno pojavila žarišča smrekovih podlubnikov. Napoved za leto 2023 smo naredili s pragom 0,30, ki pa se je izkazal za prenizkega, saj je bila kar tretjina modelskih celic lažno pozitivnih. Optimalen prag za verjetnostni model v letu 2023 je bil 0,40. Povprečen optimalen prag v obdobju 2020–2023 je bil 0,45, ki ga predlagamo za izdelavo verjetnostne napovedi v naslednjem letu.
Keywords: gozdovi, varstvo gozdov, navadna smreka, Picea abies, sanitarni posek, napoved, ogroženost, model, validacija, zmogljivost, zanesljivost, točnost, natančnost, AUC, občutljivost, specifičnost
Published in DiRROS: 11.12.2023; Views: 513; Downloads: 202
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3. Preverjanje verjetnostne in količinske napovedi sanitarnega poseka smreke zaradi podlubnikov v Sloveniji v 2022Nikica Ogris, Maarten De Groot, 2022, other scientific articles Keywords: gozdovi, zdravje gozdov, sanitarni posek, napovedi, podlubniki, Slovenija, navadna smreka, Picea abies, ogroženost, model, validacija, zmogljivost, zanesljivost, točnost, natančnost, AUC, občutljivost, specifičnost
Published in DiRROS: 14.12.2022; Views: 634; Downloads: 358
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4. Preverjanje kratkoročnih napovedi sanitarnega poseka smreke in jelke zaradi podlubnikov v Sloveniji v 2021Nikica Ogris, Maarten De Groot, 2021, other scientific articles Keywords: navadna smreka, Picea abies, bela jelka, Abies alba, sanitarni posek, napovedi, ogroženost, modeli, validacija, zmogljivost, zanesljivost, točnost, natančnost, AUC, občutljivost, specifičnost
Published in DiRROS: 14.12.2021; Views: 948; Downloads: 709
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5. Preverjanje kratkoročnih napovedi sanitarnega poseka smreke in jelke zaradi podlubnikov v Sloveniji v 2020Nikica Ogris, Maarten De Groot, 2020, other scientific articles Keywords: navadna smreka, Picea abies, bela jelka, Abies alba, sanitarni posek, napovedi, ogroženost, modeli, validacija, zmogljivost, zanesljivost, točnost, natančnost, AUC, občutljivost, specifičnost
Published in DiRROS: 14.01.2021; Views: 1233; Downloads: 1264
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