Original title: Úloha glykokonjugátů v buněčné signalizaci - CEACAMI a Fcε R1
Translated title: The role of glycoconjugates in cell signalling - CEACAM1 and Fc ε RI
Authors: Dudková, Lenka ; Šmíd, František (advisor) ; Stiborová, Marie (referee) ; Bílková, Zuzana (referee)
Document type: Doctoral theses
Year: 2006
Language: cze
Abstract: [cze] [eng]
CEACAM1 je lidský membránový glykoprotein. První části této práce se zabývá izolací CEACAM1 ze žluče, následným stanovením jeho čistoty a výtěžností použité metody. Je zde popsána imunoafinitní chromatografie CEACAM1 na hydrazidové celulose s orientovaně vázanou monoklonální protilátkou anti-CEA F34-187. Principem této metody je orientovaná vazba oxidované protilátky na aktivovaný hydrazidový matrix. Hrubý proteinový extrakt ze žluči byl aplikován na afinitní kolonu a CEACAM1 byl eluován 6 M guanidinem-HCl. Imunopozitivní zóna CEACAM1 (85 kDa) byla detekována western blotem s anti-CEA protilátkou. Pravděpodobně z důvodu značné glykosylace molekuly nebylo možné použít obvyklé metody detekce proteinů, kromě detekce s konkanavalinem AHRP. Carcinoembryonic cell adhesion molecule 1 (CEACAM1) is human membrane glycoprotein. The purpose of the first part of this thesis was to isolate CEACAM1 glycoprotein from bile and characterise its purity and recovery that have not been described before. Affinity chromatography of CEACAM1 on hydrazide-activated cellulose with immobilized monoclonal anti-CEA F34-187 antibodies is described. The method is based on the oxidation of the immunoglobulin carbohydrate component located on the Fc fragment of antibody by periodate and oriented bounding to hydrazide-activated matrix. Crude protein fraction from bile was applied onto the affinity column and CEACAM1 was eluted with 6 M guanidine-HCl. A single immunopositive 85 kDa band of CEACAM1 was detected on the western blot with anti-CEA antibody. Probably due to the high glycosylation of CEACAM1 any common method of protein staining was not applicable except staining with lectin.

Institution: Charles University Faculties (theses) (web)
Document availability information: Available in the Charles University Digital Repository.
Original record: http://hdl.handle.net/20.500.11956/7597

Permalink: http://www.nusl.cz/ntk/nusl-270081


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Universities and colleges > Public universities > Charles University > Charles University Faculties (theses)
Academic theses (ETDs) > Doctoral theses