[Purification of horseradish peroxidase conjugated IGG by affinity chromatography. Comparison of glutaraldehyde and periodate conjugation].
By affinity chromatography of the coupling mixture on Concanavalin-A-Sepharose unlabeled IgG is completely removed. HRP conjugated IgG is then separated from free HRP by gelfiltration. Glutaraldehyde conjugation yielded 20 to 68% unlabeled IgG, depending on the duration of glutaraldehyde addition, and periodate conjugation yielded 10 to 28% unlabeled IgG. By this method fractions of conjugates with 3fold spec. act. were obtained by the glutaraldehydemethod and with 1.6fold spec. act. by periodate method as compared with gelfiltration. The periodate method guarantees a 3fold higher yield of HRP-labeled IgG compares with glutaraldehyde method. Both methods produce conjugates of the same spec. act. although the denaturation of HRP is much greater at periodate conjugation. When applying of affinity chromatography and gelfiltration. Crude HRP with 7% spec. act. of purified HRP resulted in conjugates having a spec. act. of 85% of those with purified HRP.
D000818:Animals; D002846:Chromatography, Affinity; D003208:Concanavalin A; D005976:Glutaral; D006735:Horseradish Peroxidase; D007074:Immunoglobulin G / Q000302:isolation & purification*; D010504:Periodic Acid; D010544:Peroxidases; D011817:Rabbits; D012685:Sepharose
D007074:Immunoglobulin G; D010504:Periodic Acid; D003208:Concanavalin A; D012685:Sepharose; D006735:Horseradish Peroxidase; D010544:Peroxidases; D005976:Glutaral
Reinigung von Peroxidase-markiertem IgG durch Affinitätschromatographie. Vergleich von Glutaraldehyde- und Perjodatkopplung.