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Vzpostavitev avtomatizirane priprave vzorcev terapevtskih monoklonskih protiteles za kapilarno gelsko elektroforezo v prisotnosti natrijevega dodecilsulfata
ID Smolič, Katarina (Author), ID Podgornik, Aleš (Mentor) More about this mentor... This link opens in a new window, ID Mavri, Jan (Comentor)

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Abstract
Analitika terapevtskih monoklonskih protitelesa (mAb) temelji na uporabi širokega spektra analitskih metod. V našem delu smo se osredotočili na pripravo vzorcev z robotskim sistemom TECAN Freedom EVO® 200 na dveh različnih izvedbah (nereducirajoči in reducirajoči pogoji) kapilarne gelske elektroforeze v prisotnosti natrijevega dodecilsulfata (CE-SDS). Naše delo je v prvi fazi temeljilo na optimizaciji izvedbe obeh CE-SDS analiz preko določitve ustreznega načina injiciranja. V nadaljevanju smo nato ocenili uspešnost in ustreznost vzorcev pripravljenih s pomočjo robota v primerjavi s klasično ročno pripravljenimi vzorci. Za konec smo obe izvedbi CE-SDS metode dodatno optimizirali tako, da smo podaljšali rok uporabe reagentov v pladnjih za pufre. Pokazali smo, da način injiciranja nima signifikantnega vpliva na vrednosti spremljanih parametrov obeh izvedb CE-SDS analiz. Opažene razlike lahko pripišemo variabilnosti metod ali delno ročno izvedeni obdelavi rezultatov. Prav tako smo dokazali, da je avtomatizirana priprava vzorcev ekvivalentna klasični ročni pripravi vzorcev, s čimer smo tako potrdili našo glavno hipotezo. Končni rezultat testiranja je tudi potrjena stabilnost vial napolnjenih z SDS-MW gelom (shranjevanje na sobni temperaturi), kot tudi vial napolnjenih z UPW (shranjevanje v hladni sobi na 2-8°C) do enega meseca.

Language:Slovenian
Keywords:biološka zdravila, terapevtska monoklonska protitelesa, kapilarna gelska elektroforeza, avtomatizacija
Work type:Master's thesis/paper
Typology:2.09 - Master's Thesis
Organization:BF - Biotechnical Faculty
Year:2024
PID:20.500.12556/RUL-154825 This link opens in a new window
COBISS.SI-ID:189095683 This link opens in a new window
Publication date in RUL:05.03.2024
Views:691
Downloads:64
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Secondary language

Language:English
Title:Establishment of automated sample preparation for therapeutic monoclonal antibodies for capillary gel electrophoresis in the presence of sodium dodecyl sulfate
Abstract:
Analytics of therapeutic monoclonal antibody (mAb) is covered by the use of a wide range of analytical methods. In our work we focused on the preparation of samples with the TECAN Freedom EVO® 200 robotic system for the purpose of capillary gel electrophoresis in the presence of sodium dodecyl sulfate (CE-SDS) in reducing and non-reducing conditions. In the first phase of our work we evaluated the appropriate mode of sample injection (hydrodynamic and electrophoretic) for both CE-SDS methods. In the second phase we then assessed the effectiveness and comparability of samples prepared with the help of a robot compared to classical manual sample preparation. Finally, both CE-SDS methods were optimized by extending the shelf life of reagents in buffer trays where we confirmed stability of vials filled with SDS-MW gel (storage at room temperature) as well as vials filled with UPW (cold room storage at 2-8°C) for up to one month. Overall, we showed that the mode of injection has no significant impact on the values of the monitored parameters of both CE-SDS methods. The observed differences can be attributed to the variability of the methods themselves or they are the consequence of combining automatic and manual integration of electropherograms. Lastly, we have demonstrated that automated sample preparation is comparable to the classical manual sample preparation, thus confirming our main hypothesis.

Keywords:biopharmaceuticals, therapeutic monoclonal antibodies, capillary gel electrophoresis, automation

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