Glycolytic activity in extracts of young rat skin

1. 1. Data have been presented which indicate the presence of most of the glycolytic enzymes in the skin of the young rat.2. 2. A cell-free extract of this tissue will stoichiometrically convert fructose diphosphate to equimolar quantities of phosphoglyceric acid and phosphoglycerol in the presence of fluoride and trace quantities of diphosphopyridine nucleotide and inorganic phosphate under anaerobic conditions. Phosphoglycerol is formed from dihydroxyactone phosphate by a "diphosphopyridine nucleotide"-requiring glycerophosphate dehydrogenase. If P32-labeled inorganic phosphate is used, 60% of the total radioactivity is incorporated equally into phosphoglyceric acid and phosphoglycerol. This incorporation is inhibited by iodoactate.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32466/1/0000551.pd

A Sulfhydryl-Reactive Ruthenium (II) Complex and Its Conjugation to Protein G as a Universal Reagent for Fluorescent Immunoassays

To develop a fluorescent ruthenium complex for biosensing, we synthesized a novel sulfhydryl-reactive compound, 4-bromophenanthroline bis-2,2′-dipyridine Ruthenium bis (hexafluorophosphate). The synthesized Ru(II) complex was crosslinked with thiol-modified protein G to form a universal reagent for fluorescent immunoassays. The resulting Ru(II)-protein G conjugates were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The emission peak wavelength of the Ru(II)-protein G conjugate was 602 nm at the excitation of 452 nm which is similar to the spectra of the Ru(II) complex, indicating that Ru(II)-protein G conjugates still remain the same fluorescence after conjugation. To test the usefulness of the conjugate for biosensing, immunoglobulin G (IgG) binding assay was conducted. The result showed that Ru(II)-protein G conjugates were capable of binding IgG and the more cross-linkers to modify protein G, the higher conjugation efficiency. To demonstrate the feasibility of Ru(II)-protein G conjugates for fluorescent immunoassays, the detection of recombinant histidine-tagged protein using the conjugates and anti-histidine antibody was developed. The results showed that the histidine-tagged protein was successfully detected with dose-response, indicating that Ru(II)-protein G conjugate is a useful universal fluorescent reagent for quantitative immunoassays