We have purified tracer for use in the somatomedin C radioimmunoassay by hydrophobic interaction chromatography. Material showing greatest immunoreactivity binds to Octyl Sepharose CL-4B (Pharmacia) in a buffer mixture consisting of 130 mL of acetonitrile and 870 mL of 0.1 mol/L NH4HCO3, pH 7.8, but is eluted by increasing the acetonitrile content to 180 mL/L. As compared with tracer purified b...

Protein kinase C (PKC) was detected in the yeast Saccharomyces cerevisiae with bovine myelin basic protein as the phosphate acceptor. The enzyme was purified at least 500-fold by a four-step column chromatographic procedure (phenyl-Sepharose CL-4B, Mono Q, Heparin-5PW, and hydroxyapatite). The molecular mass was approximately 90 kDa, as estimated by gel-filtration analysis. Yeast PKC was activa...

Lipoxygenase, Enzyme Purification, Hydrophobic Chromatography Lipoxygenase was purified from rat liver cytosolic fraction by a method involving two successive chromatographic steps on Sephacryl S-200 and Phenyl Sepharose CL-4B. The enzyme has a molecular weight of 96 Kdal and it seems to be composed of two identical subunits. Chromatofocusing of the enzyme revealed a single band of activity at ...

Native and de novo synthesized heparan sulfate-proteoglycan (HS-PG) of basement membranes from isolated whole glomeruli were characterized. Sepharose CL-6B chromatograms of [35S]sulfate-labeled de novo synthesized HS-PG extracted from whole glomeruli indicated identical molecular weight characteristics to that isolated from purified basement membranes (Mr of intact HS-PG approximately equal to ...

We prepared two monoclonal antibodies-A-I30 and A-I4 to HDL apo A-I-with the ultimate goals of expressing and overproducing the valuable immunodiagnostic single chain Fv by phage display libraries in E, coli. Monoclonal antibodies were produced by immunizing mice with apolipoprotein A-I, and purified from ascitic fluid by affinity chromatography on a Protein A Sepharose CL-4B column. The specif...

Purified antibodies from an antiserum against S-phase proteins of the myxomycete Physarum polycephalum were attached to protein-A-Sepharose CL-4B. A late G2-phase extract that contained a mitosis-stimulating protein was applied to this immunoadsorbent, and the mitosis-stimulating protein was enriched by a factor of ten. This protein, which is present in the cell in low amounts, is synthesized i...

Human platelet proteoglycan (P.PG) was prepared from a 4 M-guanidinium chloride platelet extract in the presence of proteinase inhibitors. The purification procedure included CsCl-density-gradient centrifugation, DEAE-Sepharose CL-6B ion-exchange chromatography and f.p.l.c. on a Mono Q HR 5/5 column. P.PG was recovered as a polydisperse molecule, but the protein core appeared to be at least 90%...

This study concerned the production, purification and application of extracellular chitinase from Cellulosimicrobium cellulans strain 191. In shaken flasks the maximum yield of chitinase was 6.9 U/mL after 72 h of cultivation at 25°C and 200 rpm. In a 5 L fermenter with 1.5 vvm aeration, the highest yield obtained was 4.19 U/mL after 168 h of fermentation at 25°C and 200 rpm, and using 3 vvm, i...

Cibacron Blue 3G-A (CB3G-A, I) was investigated as a ternary complex analogue of lactate dehydrogenase and phosphoglycerate kinase as had been suggested earlier (Stellwagen, E. (1977) Accts. Chem. Res. 10, 92-98). CB3G-A and Procion Brilliant Blue (PBB, II), a structural isomer of the Cibacron dye without the sulfonated benzoyl moiety, were attached covalently to Sepharose CL-6B. The two enzyme...

An extreme thermophile that produces a heat-stable proteinase was isolated from hot-spring water and classified as Thermomicrobium sp. KN-22 (growth temperature, 50-83 degrees C; and optimum growth temperature, 70 degrees C). The proteinase was purified from the culture broth of this strain by fractionation with ammonium sulfate, chromatography on columns of DEAE-cellulose and CM-Sepharose CL-6...