Proteomics-based approaches for validation of antibodies against OTR and V1aR

master's thesis

• Title (croatian): Validacija antitijela za OTR i V1aR pristupima temeljenima na proteomici
• Author: Bojan Kuridža
• Mentor: Markus Muttenthaler (mentor)
• Committee member: Marko Močibob (predsjednik povjerenstva)
• Committee member: Ivan Kodrin (član povjerenstva)
• Committee member: Iva Juranović Cindrić (član povjerenstva)
• Committee member: Đani Škalamera (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2023-06-06, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry

Abstract

The oxytocin receptor (OTR) and the vasopressin 1a receptor (V1aR) are two G protein-coupled
receptors involved in mediating various physiological processes and social and sexual
behaviors. Producing specific antibodies to study these receptors is challenging due to their
sequence homology and structure similarity, along with two additional closely related
vasopressin receptors. Thus, this thesis aimed to validate ten antibodies to OTR and seven
antibodies to V1aR produced against distinctive receptor peptide epitopes, using western blot
and immunoprecipitation followed by mass spectrometry. The validation process pointed
towards one V1aR antibody (1G1), which showed promising results in both western blot and
immunoprecipitation experiments, and needs to be further tested. The best OTR antibody
identified in immunoprecipitation experiments (1B2) did not perform well in western blots.

Abstract (croatian)

Oksitocinski receptor (OTR) i vazopresinski 1a receptor (V1aR) dva su receptora spregnuta sa
G-proteinom uključena u posredovanje raznih fizioloških procesa te društvenog i seksualnog
ponašanja. Proizvodnja specifičnih antitijela za proučavanje ovih receptora predstavlja izazov
zbog sličnosti njihovog slijeda i strukture, kao i sličnosti dva dodatna blisko povezana
vazopresinska receptora. Stoga, cilj je ovog rada bio validirati deset antitijela za OTR i sedam
antitijela za V1aR proizvedenih prema karakterističnim peptidnim epitopima receptora,
koristeći western blot i imunoprecipitaciju povezanu s masenom spektrometrijom. Jedno
antitijelo za V1aR (1G1) u procesu validacije pokazalo je obećavajuće rezultate u
eksperimentima western blota i imunoprecipitacije. Najbolje od OTR antitijela identificiranih
u eksperimentima imunoprecipitacije (1B2) nije se pokazalo uspješnim u western blotovima.

• Keywords: antibody validation; G protein-coupled receptor; immunoprecipitation; mass spectrometry; oxytocin receptor; vasopressin 1a receptor; western blot
• Keywords (english): imunoprecipitacija; masena spektrometrija; oksitocinski receptor; receptor spregnut sa G-proteinom; validacija antitijela; vazopresinski 1a receptor; western blot
• Language: english
• URN:NBN: urn:nbn:hr:217:185973
• 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: 2023-06-09 10:18:20