Preparation of silica coated cobalt ferrite magnetic nanoparticles for the purification of histidine-tagged proteins


Thesis Type: Postgraduate

Institution Of The Thesis: Orta Doğu Teknik Üniversitesi, Faculty of Arts and Sciences, Department of Chemistry, Turkey

Approval Date: 2011

Student: GÜLFEM AYGAR

Co-Supervisor: NECATİ ÖZKAN, MÜRVET VOLKAN

Abstract:

The magnetic separation approach has several advantages compared with conventional separation methods; it can be performed directly in crude samples containing suspended solid materials without pretreatment, and can easily isolate some biomolecules from aqueous systems in the presence of magnetic gradient fields. This thesis focused on the development of new class of magnetic separation material particularly useful for the separation of histidine-tagged proteins from the complex matrixes through the use of imidazole side chains of histidine molecules. For that reason surface modified cobalt ferrite nanoparticles which contain Ni-NTA affinity group were synthesized. Firstly, cobalt ferrite nanoparticles with a narrow size distribution were prepared in aqueous solution using the controlled coprecipitation method. In order to obtain small size of agglomerates two different dispersants, oleic acid and sodium chloride, were tried. After obtaining the best dispersant and optimum experimental conditions, ultrasonic bath was used in order to decrease the size of agglomerates. Then, they were coated with silica and this was followed by surface modification of these nanoparticles by amine in order to add functional groups on silica shell. Next, –COOH functional groups were added to silica coated cobalt ferrite magnetic nanoparticles through the NH2 groups. After that Nα,Nα-Bis(carboxymethyl)-L-lysine hydrate, NTA, was attached to carboxyl side of the structure. Finally, nanoparticles were labeled with Ni (II) ions. The size of the magnetic nanoparticles and their agglomerates were determined by FE-SEM images, particle size analyzer, and zeta potential analyzer (zeta-sizer). Vibrational sample magnetometer (VSM) was used to measure the magnetic behavior of cobalt ferrite and silica coated cobalt ferrite magnetic nanoparticles. Surface modifications of magnetic nanoparticles were followed by FT-IR measurements. ICP-OES was used to find the amount of Ni (II) ion concentration that was attached to the magnetic nanoparticle.