20.500.12556/DKUM-89663 Optimizacija in vitro stimulacije limfocitov s fitohemaglutininom (PHA) in lipopolisaharidi (LPS) diplomsko delo univerzitetnega študijskega programa I. stopnje Optimisation of in vitro stimulation of lymphocytes with phytohaemagglutinin (PHA) and lipopolysaccharides (LPS) Stimulacija limfocitov s fitohemaglutininom(PHA) in lipopolisaharidi(LPS) je ključna metoda za preučevanje imunskih odzivov v eksperimentalnih pogojih. Optimizacija koncentracij omenjenih imunostimulatorjev je bistvena za zagotavljanje fiziološko relevantnih in natančnih rezultatov. V diplomski nalogi smo preučevali odziv limfocitov na različne koncentracije PHA in LPS, ki inducirajo 70% maksimalnega odziva. Pri tem smo prav tako želeli ugotoviti, če najvišje koncentracije PHA in LPS povzročijo celično smrt. Limfocite smo izolirali iz mononuklearnih celic periferne krvi(PBMC) zdravih darovalcev in jih gojili v prisotnosti različnih koncentracij PHA in LPS. Proliferacijo celic smo merili s pomočjo barvila alamarBlue in določili celično viabilnost z uporabo pretočne citometrije. Rezultati so pokazali, da je PHA pri vmesni koncentraciji 1-2,5 μg/mL učinkovit stimulator za doseg 70% maksimalnega odziva limfocitov, hkrati pa najvišje koncentracije PHA povzročajo celično smrt. Nasprotno je bil LPS neuspešen zaradi velike variabilnosti med donorji. Naše ugotovitve prispevajo k boljši standardizaciji eksperimentalnih protokolov pri imunoloških raziskavah. Lymphocyte stimulation with phytohaemagglutinin(PHA) and lipopolysaccharides(LPS) is a key method for studying immune responses under experimental conditions. Optimisation of the concentrations of these immunostimulants is essential to ensure physiologically relevant and accurate results.We studied the response of lymphocytes to different concentrations of PHA and LPS, which induced 70% of the maximum response. We also wanted to determine if high concentrations of PHA and LPS induce cell death. Lymphocytes were isolated from peripheral blood mononuclear cells (PBMCs) of healthy donors and cultured in the presence of different concentrations of PHA and LPS. We measured cell proliferation with alamarBlue and determined cell viability using flow cytometry. Our results demonstrated that PHA, at intermediate concentrations of 1-2.5 μg/mL, effectively stimulates a 70% of the maximum lymphocyte response, while at the same time the highest PHA concentrations induce cell death. In contrast, LPS was unsuccessful due to high inter-donor variability. Our findings contribute to better standardisation of experimental protocols in immunology research. limfociti stimulacija pretočna citometrija PHA LPS optimizacija lymphocytes stimulation flow cytometry PHA LPS optimisation true true false [P. Počivavšek] Slovenski jezik Angleški jezik Diplomsko delo/naloga 2024-08-01 21:45:19 2024-09-19 12:50:25 2024-09-28 03:20:47 0000-00-00 00:00:00 2024 0 Maribor 0 Maribor 1 spletni vir (1 datoteka PDF (VIII, 28 f.)) 0000-00-00 NiDoloceno NiDoloceno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 616-097:612.112(043.2) 209245699 UN_Pocivavsek_Patricija_2024.pdf UN_Pocivavsek_Patricija_2024.pdf 1 F2F280E1085DB408081E1FB8129AE502 2a01b911fe02250ceb5bbff29c461c47a86b52107c9719bae0cc7326fbe08365 a753b16f-7667-11ef-aec7-00155d000105 https://dk.um.si/Dokument.php?lang=slv&id=185463 Fakulteta za kemijo in kemijsko tehnologijo 0 0 0