DARWIN Digitale Dissertationen German Version Strich

FU Berlin
Digitale Dissertation

Sigrun Lerch :
Investigation of the antigenic protein fractions of a Sarcoptes mite extract by SDS-PAGE, two dimensional electrophoresis and sequenz analysis
Untersuchung der antigenen Proteinfraktionen eines Sarcoptes-Milben-Extraktes durch SDS-PAGE, Immunoblot, Zweidimensionale Gelelektrophorese und Sequenzanalyse

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Abstract

This research project was aimed to establish wich fractions of the complex protein solution of the Sarcoptes mite extract would be able to bind specific Sarcoptes antibodies and would thus be effective in an antigenic fashion.
For the succes of this experimental study a qualitatively and quantitatively suficient antigen preparation of mite extract solution was indispensible.
To achieve this objective, an extract of Sarcoptes scabiei var. vulpes was prepared by harvesting mites collected from crusts of carcasses of naturally infested red foxes. The mites were subsequently isolated by a migration technique and then processed.
The protein mixture was produced through homogenization of the entire mitebody also defined as "raw antigen".
By applying the one dimensional SDS-PAGE the proten mixture could be separated into 33 bands in the range of 15-225 kDa.
18 of these protein bands were classified and bound by specific Sarcoptes antibodies in the immunoblot.
By using the technique of Two-dimensional electrophoresis, the proteins were arranged according to their distinct isoelectric points as well as their variable molecular weights. Thus it was possible to present the specific antibody binding protein bands in greater detail. Additionally, by applying this method the greatest biochemical purity of the protein spots could be achieved. This made the posterior sequenz of operation feasible.
Before undertaking the sequenz analysis the protein spots were blotted on a PVDF membrane and differentiated by Coomassie coloration. In the applied sequencing process 2 out of 8 spots yielded results.
The spots showed the following sequences:
Spot 11: (Als/Ser) Pro Asn His Asp Lys Ala
Phe Asp (Val/Glu) Leu - Val
Spot 12: (Met/Gln) (Lys/Ser/Gly) Asn
(Lys/Ser) Ala Val Leu Leu Gly Val
Tyr Glu Asn - Asp

Table of Contents

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Titelblatt, Inhaltsverzeichnis, Lebenslauf
1. Einleitung
2. Literaturübersicht
3. Material und Methoden
4. Ergebnisse
5. Diskussion
6. Zusammenfassung
7. Summary
8. Literaturverzeichnis

More Information:

Online available: http://www.diss.fu-berlin.de/2001/285/indexe.html
Language of PhDThesis: german
Keywords: Protein Analysis; Sarcoptes Scabiei; SDS-PAGE
DNB-Sachgruppe: 34 Veterinärmedizin
Date of disputation: 16-Jul-2001
PhDThesis from: Fachbereich Veterinärmedizin, Freie Universität Berlin
First Referee: Univ.-Prof. Dr. Eberhard Schein
Second Referee: Univ.-Prof. Dr. Michael F. G. Schmidt
Third Referee: Univ.- Prof. Dr. Dieter Ebner
Contact (Advisor): sigrun-lerch@gmx.de
Date created:03-Feb-2002
Date available:04-Feb-2002

 

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