title: Purification of sheep immunoglobin G using protein A trapped in sol-gel glass.
abstract: An effective method for purifying immunoglobulins has been developed utilizing a sol-gel glass support system. Sol-gel glass is an effective support for chemically active ligands entrapped in this medium at room temperature. There are two problems associated with the utilization of such sol-gel glasses for entrapping macromolecules. One is the phenomenon of nonspecific absorption of proteins onto the glass. The second is that only a portion of the entrapped molecules may retain their biological activity. In the present study a sol-gel glass was treated with gamma-aminopropyltriethoxysilane to provide a matrix that eliminated nonspecific absorption of proteins. The method of entrapping molecules was modified to increase the proportion of entrapped molecules that retained their reactivity. Protein A was entrapped in the modified sol-gel glass column and used to purify IgG from sheep sera by affinity chromatography. The purity of the IgG, as determined by SDS-PAGE, was comparable to that obtained from commercially available protein A columns and, if the capacity of the column was not exceeded, the yield approached 100%. Although the quality and quantity of the yields were comparable, the methodology described herein can be accomplished more rapidly and with greater ease. Furthermore, the sol-gel glass ligand preparation is extremely stable and can be reused for isolation of gamma-globulin. The technique has great potential for isolating macromolecules utilizing various ligands.
authors.affiliation: Department of Pediatrics, Jefferson Medical School, Thomas Jefferson University, Wilmington, Delaware.
authors.forename: R
authors.initials: R
authors.lastname: Zusman
authors.forename: D A
authors.initials: DA
authors.lastname: Beckman
authors.forename: I
authors.initials: I
authors.lastname: Zusman
authors.forename: R L
authors.initials: RL
authors.lastname: Brent
mesh_terms: D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D005782:Gels; D005898:Glass; D007074:Immunoglobulin G / Q000302:isolation & purification*; D008722:Methods; D012756:Sheep; D013205:Staphylococcal Protein A
chemical_list: D005782:Gels; D007074:Immunoglobulin G; D013205:Staphylococcal Protein A