Optimizacija visokoprotočnih metoda za analizu N-glikozilacije mišjeg imunoglobulina G

master's thesis

• Title (english): Optimisation of high-throughput analysis of mouse immunoglobulin G N-glycosylation
• Author: Mia Mrčela
• Mentor: Gordan Lauc (mentor)
• Committee member: Marko Močibob (predsjednik povjerenstva)
• Committee member: Sanda Rončević (član povjerenstva)
• Committee member: Biserka Prugovečki (član povjerenstva)
• Committee member: Mirta Rubčić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2017-05-31, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry

Abstract

N-glikozilacija ima strukturnu ulogu te utječe na efektorske funkcije imunoglobulina G (IgG-a). Promjena glikozilacije povezuje se s kompleksnim bolestima za čiji se mehanizam nastanka kao modelni organizmi koriste miševi. Miševi s uklonjenim genima (eng. knockout) za receptore specifične za fragment koji kristalizira (FcγR) mogu dati uvid kako N-glikozilacija utječe na interakcije fragmenta koji kristalizira (Fc) i receptora Fcγ u reakcijama imunološkog sustava. Potrebno je razviti osjetljive visokoprotočne metode kojima se osim velikog broja uzoraka, može analizirati i N-glikozilacija manjih koncentracija IgG-a kao što je mišji IgG. Metodom LC-MS analizirana je N-glikozilacija IgG-a izoliranog iz seruma divljeg tipa i miševa s uklonjenim genima koji kodiraju FcγRI, FcγRIIB, FcγRIII, FcγRIV te γ teški lanac fragmenta Fc. Također, uspoređeni su glikanski profili IgG-a iz miševa divljeg tipa te miševa s uklonjenim genima za receptore Fcγ i γ lanac. Djelomično je optimizirana metoda CGE-LIF kojom je analizirana N-glikozilacija ljudskog IgG-a.

Abstract (english)

N-glycosylation has structural roll and modulate immunoglobulin G (IgG) effector functions. Glycosylation changes are associated with complex diseases and mouse model organism is used for studies of disease development mechanism. Fragment crystallizable γ receptor (FcγR) knockout mouse can provide insight into how IgG N-glycosylation affects the fragment crystallizable (Fc) and FcγR interaction during immune reaction. It is necessary to bring into practice sensitive high-throughput methods not only for large number of samples, but also for N-glycosylation analysis in samples with low IgG concentrations as for example mouse IgG. N-glycosylation of FcγRI, FcγRIIB, FcγRIII, FcγRIV and γ-chain knockout mouse IgG is analysed by LC-MS method. Furthermore, glycoprofiles of IgG glycoforms from wild type mouse strains are compared with FcγR and γ chain knockouts. CGE-LIF method is partially optimised and human IgG N-glycosylation is analysed.

• Keywords: imunoglobulin G; N-glikozilacija; receptori Fcγ; tekućinska kromatografija spregnuta sa spektrometrijom masa; kapilarna gel elektroforeza
• Keywords (english): N-glycosylation; immunoglobulin G; Fcγ receptors; liquid chromatography coupled with mass spectromety; capillary gel electrophoresis
• Language: croatian
• URN:NBN: urn:nbn:hr:217:727906
• Study programme: Title: Graduate university studies in chemistry; specializations in: Research programme; Course: Research programme; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra kemije (magistar/magistra kemije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-06-05 13:50:42