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master's thesis
Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies
2020. urn:nbn:hr:193:491332
Žarković, Nina
University of Rijeka
Faculty of Biotechnology and Drug Development

Cite this document

Žarković, N. (2020). Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies (Master's thesis). Rijeka: University of Rijeka. Retrieved from https://urn.nsk.hr/urn:nbn:hr:193:491332

Žarković, Nina. "Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies." Master's thesis, University of Rijeka, 2020. https://urn.nsk.hr/urn:nbn:hr:193:491332

Žarković, Nina. "Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies." Master's thesis, University of Rijeka, 2020. https://urn.nsk.hr/urn:nbn:hr:193:491332

Žarković, N. (2020). 'Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies', Master's thesis, University of Rijeka, accessed 27 April 2024, https://urn.nsk.hr/urn:nbn:hr:193:491332

Žarković N. Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies [Master's thesis]. Rijeka: University of Rijeka; 2020 [cited 2024 April 27] Available at: https://urn.nsk.hr/urn:nbn:hr:193:491332

N. Žarković, "Cell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies", Master's thesis, University of Rijeka, Rijeka, 2020. Available at: https://urn.nsk.hr/urn:nbn:hr:193:491332

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Metadata
TitleCell-type specific glycosylation patterns of mouse CD63 detected by newly produced mouse monoclonal antibodies
Title (croatian)Stanično specifične glikozilacije proteina CD63 otkrivene novoproizvedenim mišjim monoklonskim protutijelima
AuthorNina Žarković
MentorKristina Grabušić (mentor)
MentorMladenka Malenica Staver (komentor)
Committee memberElitza Petkova Markova - Car (predsjednik povjerenstva)
Committee memberIlija Brizić (član povjerenstva)
GranterUniversity of Rijeka
(Faculty of Biotechnology and Drug Development)
Rijeka
Defense date and country2020-07-23, Croatia
Scientific / art field,
discipline and subdiscipline		BIOTECHNICAL SCIENCES
Biotechnology
Abstract
CD63 is an evolutionary conserved transmembrane glycoprotein which is ubiquitously expressed in mammalian cells and abundantly located in late endosomes and lysosomes. It has no enzymatic activity, but exerts its functions by interacting with other proteins to act on a wide range of cellular processes like cytoskeletal organisation, cell adhesion and motility. At least some of these CD63 functions might explain the association of CD63 with cancer metastasis seen in clinical settings. However, for a deeper understanding of CD63 biology, the research community requires novel antibodies suitable for a systematic biochemical characterisation of CD63. This study provides characterisation of two newly produced mouse monoclonal antibodies directed against mouse CD63: mCD63.07 and mCD63.08. Both antibodies bind to a linear epitope located at the extracellular portion of CD63 and are highly specific and sensitive in CD63 detection by immunoblot and immunofluorescence analyses based on signal comparison between wild type and CD63 knockout mouse fibroblasts. Moreover, the knockout validated antibodies were able to distinguish multiple CD63 glycoforms in several murine cell lines of different origins indicating that CD63 might have cell-type specific forms and functions. Finally, with the newly developed immunofluorescence protocol including protein denaturation, reduction and deglycosylation, novel antibodies detected significant CD63 amounts in cellular parts other than late endosomes and lysosomes.Taken together, the newly developed antibodies might become a valuable tool to study CD63 biology thanks so the their two main features: ability to detect multiple CD63 glycoforms and to identify potentially novel cellular localisation of CD63.
Abstract (croatian)
Tetraspanin CD63 je ubikvitarno izražen protein u sisavaca koji se većinom nalazi u kasnim endosomima i lizosomima. CD63 nije enzimski aktivan, on svoje mnogobrojne funkcije izvršava preko interakcija s drugim proteinima te pritom sudjeluje u širokom rasponu staničnih procesa poput citoskeletne organizacije, stanične adhezije i pokretljivosti. U kliničkim je postavkama uspostavljena veza između CD63 i metastatskog potencijala tumora, za koju mogu biti odgovorne gore spomenute funkcije CD63. Međutim, za potpunije razumijevanje biologije CD63 proteina, istraživačkoj su zajednici potrebna nova protutijela pogodna za sustavnu biokemijsku karakterizaciju CD63 proteina. Ovim su radom karakterizirana dva novoproizvedena mišja monoklonska protutijela usmjerena protiv mišjeg CD63: mCD63.07 i mCD63.08. Na temelju usporedbe signala između mišjih fibroblasta divljeg tipa i mišjih fibroblasta CD63 knockout tipa, utvrđeno je da se oba protutijela vežu na linearni epitop smješten na izvanstaničnom dijelu CD63 i da su vrlo specifična i osjetljiva u detekciji CD63 imunofluorescentnom i imunoblot metodom. Nadalje, knockout validirana protutijela mogu razlikovati višestruke glikoforme CD63 u nekoliko mišjih staničnih linija različitog podrijetla, što ukazuje da CD63 može imati stanično specifične forme i funkcije. Naposlijetku, s novorazvijenim protokolom imunofluorescencije, koji uključuje denaturaciju proteina, redukciju i deglikozilaciju, nova protutijela detektirala su značajne količine CD63 u odjeljcima stanice koji nisu kasni endosomi i lizosomi. Na temelju dobivenih rezultata, novorazvijena antitijela mogla bi postati vrijedan alat za proučavanje biologije CD63 zahvaljujući njihovim dvjema glavnim značajkama: sposobnosti otkrivanja više glikoformi CD63 i identificiranja potencijalno nove stanične lokalizacije CD63.
Keywords
Keywords (croatian)
Languageenglish
URN:NBNurn:nbn:hr:193:491332
Study programmeTitle: Biotechnology in medicine
Study programme type: university
Study level: graduate
Academic / professional title: magistar/magistra biotehnologije u medicini (magistar/magistra biotehnologije u medicini)
Type of resourceText
File originBorn digital
Access conditionsOpen access
Embargo expiration date: 2022-07-24
Terms of useLicence In copyright icon
Created on2020-07-24 15:51:18