Assay and Preparation

Harlow, E. C. Weinbaeh, and L. 8. Diamond, Comp. Biochem. Biophya. (in press). 

Kramer and F. Salvenmoser, Hoppe-Seyler s Z. Physiol. Chem. 346, 310 (1966). 

Recently, Rydstrom et al. (107) demonstrated that transhydroge-nase was solubilized efficiently and selectively from beef heart sonic sub-mitochondrial particles by lysolecithin, giving a preparation that was about seven times as active as particles. In contrast to earlier preparations this lysolecithin extract was devoid of all cytochromes except cytochrome c. 

Assay of AB-speciflc transhydrogenase is principally identical to that of BB-speciflc transhydrogenase (see Section II,C) i.e., with the natural nicotinamide nucleotides, an enzymic regenerating system is used to keep the concentration of one of the substrates constant (30, 32, 66-68, 66) 

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Volume II. Preparation and Assay of Enzymes Edited by Sidney P. Colowick and Nathan O. Kaplan... 

Volume VI. Preparation and Assay of Enzymes (Continued) Preparation and Assay of Substrates Special Techniques... 

The zero standard is prepared and assayed, but is not included in calibration curve regression. 

O Neill and coworkers have also sought to address the problem of the metabolically susceptible CIO acetal linkage . A series of CIO carba dimers were prepared and assayed for antitumour activity. The two most potent compounds that were prepared are two phosphate ester finked dimers 115 and 116 (Scheme 40). They are principally active against leukaemia, colon and certain melanoma and breast cancer cell lines in the NCI 60-cell line assay. 

Procedure - Equilibrate the Standard Preparation and the Assay Preparation in the dark at 80° for 2.5 hours, accurately timed. Cool, and introduce equal volumes (5 to 10 1) of the heat-equilibrated Standard Preparation and Assay Preparation into the high-pressure liquid chromatograph (See chromatography <621>) by means of a suitable sampling valve. Measure the peak responses obtained for the Assay Preparation and the Standard Preparation, and calculate the quantity, in mg, of C27H44O in the portion of Cholecalciferol taken by the formula 0.25C(Ay/Ae)> in which C is the concentration, in g per ml, of USP Cholecalciferol RS in the Standard Preparation, and Ay and As are the peak responses for cholecalciferol obtained for the Assay Preparation and the Standard Preparation, respectively. 

The cyanide ion in inorganic cyanides can be present as both complexed and free cyanide. In order to study the chromatography of metal cyanides, Rocklin and Johnson [48] prepared and assayed solutions of cadmium, zinc, copper, nickel, gold, iron and cobalt cyanides. Table 2.6 lists the percentage of total cyanide detected. 

We thank the British Technology Group and the Association of Anaesthetists for sponsoring the University Research Program, Professor G. Smith and Dr. C. Hanning of Leicester University, Department of Anaesthesia, for their joint interest and clinical results, and our laboratory technicians P. Keating and R. Cameron for careful preparations and assays. 

In the ceruloplasmin experiments appropriate aliquots of specimens were placed in small test tubes and assayed for Cu64 activity in a well-type scintillation counter. Analyses were made with reference to a Cu64 standard prepared and assayed at the outset and whenever other samples were assayed. This permitted correction for radioactivity decay of this 12.8-hour half-life isotope. 

Thompson, G. A. and Kapoulas, V. M. (1968) Preparation and assay of glyceryl ethers, in Methods in Enzymology, Vol. 14, J. Lowenstein, editor, Academic Press, New York, pp. 668-678. 

Standard Preparation and Assay Preparation Proceed as directed under Assay. 

Procedure Determine the peak area in the chromatograms for the Standard Preparation and Assay Preparation by separately injecting 20 iL of each into the chromatograph. Calculate the quantity in percent of sodium methyl sulfate in the sample by the formula... 

Sizer, I. W., and Jenkins, W. T. (1962). Glutamic Aspartic Transaminase from Pig Ventricles Preparation and Assay of Enzymes. Methods Enzymol 5 677. Stryer, L. (1995). Amino Acid Degradation and the Urea Cycle. In Biochemistry, 4th ed. New York Freeman. 

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