Horseradish peroxidase and microperoxidase. Their purity and binding to serum proteins.
Buffer solutions of commercially available batches of the electron microscopic enzyme tracers, horseradish peroxidase (HRP, mol wt = 40,000) and microperoxidase (MP, mol wt = 2,000), were investigated by gel filtration. The elution profiles of the HRP and MP solutions together with estimates of the peroxidatic activity of the collected fractions indicated that HRP was homogeneous with respect to molecular weight, while MP contained high molecular weight impurities with peroxidatic activity. Serum samples from four different species of tracer-injected animals (hagfish, lamprey, frog, mouse) and from control animals were investigated with the same techniques. No binding between HRP and the serum proteins or degradation of HRP was detected; whereas, MP circulated predominantly bound to serum proteins in all the investigated animals. The number average molecular weights of aqueous solutions of HRP and MP determined by vapor osmometry were 9,000 and 1,000, respectively. The results emphasize that protein binding should be evaluated when MP is used as an electron microscopic tracer. In addition, the finding that HRP and MP solutions have a much higher osmotic activity than the molecular weights of the tracers would suggest should be taken into consideration, especially when bolus injections into animals are performed.
D000818:Animals; D001798:Blood Proteins / Q000378:metabolism*; D002850:Chromatography, Gel; D006196:Hagfishes; D006420:Hemeproteins / Q000302:isolation & purification / Q000378:metabolism*; D006735:Horseradish Peroxidase / Q000302:isolation & purification / Q000378:metabolism*; D007798:Lampreys; D051379:Mice; D008970:Molecular Weight; D010544:Peroxidases / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D011896:Rana temporaria
D001798:Blood Proteins; D006420:Hemeproteins; D006735:Horseradish Peroxidase; D010544:Peroxidases; C021799:microperoxidase