Gelfiltrationchromatography-KSU：凝胶过滤色谱-堪萨斯州立大学.pptVIP

Gel filtration chromatography /legacy/college/voet/0471214957/animated_figures/ch05/f5-6.html Gel filtration (chromatography), is also known as molecular sieve chromatography. Gel filtration chromatography separates molecules according to their size and shape. The stationary phase consists of beads containing pores that span a relatively narrow size range. Smaller molecules spend more time inside the beads than larger molecules and therefore elute later (after a larger volume of mobile phase has passed through the column). Types of gels used The gels used as molecular sieves are cross linked polymers. They are uncharged and inert i.e. don’t bind or react with the materials being analyzed. Three types of gels are used: Types of gels cont… Dextran: is a homopolysaccharide of glucose residues. it’s prepared with various degrees of cross-linking to control pore size. It’s bought as dry beads, the beads swell when water is added. The trade name is sephadex. It’s mainly used for separation of small peptides and globular proteins with small to average molecular mass. Types of gels cont… 2. Polyacrylamide:these gels are prepared by cross linking acrylamide with N,N-methylene bis acrylamide. The pore size is determined by the degree of cross-linking. The separation properties of polyacrylamide gels are mainly the same as those of dextrans. They are sold as bio-gel P. They are available in wide range of pore sizes. Types of gels cont… 3. Agarose: linear polymers of D-galactose and 3,6 anhydro-1-galactose. It forms a gel that’s held together with H bonds. It’s dissolved in boiling water and forms a gel when it’s cold. The concentration of the material in the gel determines the pore size. The pores of agarose gel are much larger than those of sephadex or bio-gel p. It’s useful for analysis or separation of large globular proteins or long linear molecules such as DNA The gel filtration material that will be used in the experiment below is called Sephadex G-75 and it will s