Selection and Production of Recombinant Binders for Use in Protein Microarrays

Nonfiction, Health & Well Being, Medical, Medical Science, Microbiology
Cover of the book Selection and Production of Recombinant Binders for Use in Protein Microarrays by Mark Laible, GRIN Verlag
View on Amazon View on AbeBooks View on Kobo View on B.Depository View on eBay View on Walmart
Author: Mark Laible ISBN: 9783640994267
Publisher: GRIN Verlag Publication: August 29, 2011
Imprint: GRIN Verlag Language: English
Author: Mark Laible
ISBN: 9783640994267
Publisher: GRIN Verlag
Publication: August 29, 2011
Imprint: GRIN Verlag
Language: English

Diploma Thesis from the year 2009 in the subject Biology - Micro- and Molecular Biology, grade: 1,1, German Center of Cancer Research Heidelberg, language: English, abstract: Antibodys are essential everyday tools for molecular biologists and much depends on their performance and availabilty. Large numbers of antibodys are available already, however, these cover only a small part of the known proteom. The generation of antibodies for proteom-wide studies using conventional methods, which are based on animal immunisation, would be hardly feasible and not sustainable. Recombinant binders may be able to fill the gap of missing affinity reagents. The main advantages of recombinant binders are (1) the wide availabilty in form of fully synthetic DNA libraries (2) the independance on animal immunisation, which still is the main source of antibodies today and (3) the possibility to design completely new classes of synthetic binders with definded properties. The aim of this work was to establish the selection and purification of recombinant binders for use in protein microarrays. Binders were selected from a synthetic library of single chain fragment variable (scFv) antibodies using phage display as tool for selection. A number of binders against gamma globulins and BSA-FITC could be selected from the library and were subsequently used for establishment of downstream applications. Several methods for expression and purification of the binders were tested mostly in regard to scFv yield. Methods for detection were evaluated for their sensitivity and specificity. As the long term aim was the setup of a protein microarray based on recombinant binders, multiple immobilisation strategies were evaluated, including a one step on-chip purification and immobilisation of scFvs from crude expression samples. The on-chip purification was shown to work. However, the best results regarding signal intensity were obtained using biotinylated protein-A purified scFvs immobilised in a directed manner on streptavidin-coated glass slides. Undirected immobilisation of purified scFvs on epoxy-silane slides was as well shown to be a possibility. In a side project, the novel phage RDA (representational difference analysis) technique for PCR and DNA hybridisation based selection of binders was applied. The major focus was put onto optimizing the critical steps of amplification of the scFv library by PCR.

View on Amazon View on AbeBooks View on Kobo View on B.Depository View on eBay View on Walmart

Diploma Thesis from the year 2009 in the subject Biology - Micro- and Molecular Biology, grade: 1,1, German Center of Cancer Research Heidelberg, language: English, abstract: Antibodys are essential everyday tools for molecular biologists and much depends on their performance and availabilty. Large numbers of antibodys are available already, however, these cover only a small part of the known proteom. The generation of antibodies for proteom-wide studies using conventional methods, which are based on animal immunisation, would be hardly feasible and not sustainable. Recombinant binders may be able to fill the gap of missing affinity reagents. The main advantages of recombinant binders are (1) the wide availabilty in form of fully synthetic DNA libraries (2) the independance on animal immunisation, which still is the main source of antibodies today and (3) the possibility to design completely new classes of synthetic binders with definded properties. The aim of this work was to establish the selection and purification of recombinant binders for use in protein microarrays. Binders were selected from a synthetic library of single chain fragment variable (scFv) antibodies using phage display as tool for selection. A number of binders against gamma globulins and BSA-FITC could be selected from the library and were subsequently used for establishment of downstream applications. Several methods for expression and purification of the binders were tested mostly in regard to scFv yield. Methods for detection were evaluated for their sensitivity and specificity. As the long term aim was the setup of a protein microarray based on recombinant binders, multiple immobilisation strategies were evaluated, including a one step on-chip purification and immobilisation of scFvs from crude expression samples. The on-chip purification was shown to work. However, the best results regarding signal intensity were obtained using biotinylated protein-A purified scFvs immobilised in a directed manner on streptavidin-coated glass slides. Undirected immobilisation of purified scFvs on epoxy-silane slides was as well shown to be a possibility. In a side project, the novel phage RDA (representational difference analysis) technique for PCR and DNA hybridisation based selection of binders was applied. The major focus was put onto optimizing the critical steps of amplification of the scFv library by PCR.

More books from GRIN Verlag

Cover of the book Kritische Analyse der Budgetierung, des Better Budgeting und des Beyond Budgeting by Mark Laible
Cover of the book Gläubigerschutz durch vertragliche Abreden - financial covenants by Mark Laible
Cover of the book Gutachterliche Prüfung von 4 Fällen, angelehnt an den Münzhändlerfall, Einsturz einer Eissporthalle, Unterlassene Hilfeleistung eines Polizisten und Fahrlässige Tötung durch einen Autounfall by Mark Laible
Cover of the book Teilnehmende versus Dichte Beobachtung. Weiterentwicklung oder Konkurrenz? by Mark Laible
Cover of the book Luhmannsche Systemtheorie exemplifiziert am Casino Hohensyburg by Mark Laible
Cover of the book Les danses des sociétés créoles by Mark Laible
Cover of the book Erlernte Hilflosigkeit by Mark Laible
Cover of the book Semiotics of Law: Intent and Preparation with in a Semiosis Cycle by Mark Laible
Cover of the book Die Republik der Vereinigten Niederlande und der Jülich-Klevische Erbfolgestreit by Mark Laible
Cover of the book Normales Altern unnormal? Menschen mit geistiger Behinderung im Ruhestand by Mark Laible
Cover of the book Lernen an Stationen - Eine Möglichkeit kindgerechten Lernens by Mark Laible
Cover of the book Die kunstgeschichtliche Hermeneutik nach Oskar Bätschmann. Ein Vergleich mit dem dreistufigen Interpretationsschema von Erwin Panofsky by Mark Laible
Cover of the book Der Volksnarr und Schwankheld Nasreddin Hoca. Zur Identitätsfrage eines beliebten Protagonisten humoristischer Kurzprosa im islamischen Orient by Mark Laible
Cover of the book Softwaredokumentation im Projekt CASSAC by Mark Laible
Cover of the book Untersuchung von Unterschieden zwischen Eltern und Kinderlosen in Bezug auf diverse Einstellungen by Mark Laible
We use our own "cookies" and third party cookies to improve services and to see statistical information. By using this website, you agree to our Privacy Policy