By D.B. Roodyn (Eds.)

ISBN-10: 0080887732

ISBN-13: 9780080887739

ISBN-10: 0720442060

ISBN-13: 9780720442069

Within the face of fast advancements in automatic ideas, this e-book provides a good consultant to offer developments. It presents info on: rules and terminology of enzyme automation; automated tools illustrated via the Technicon approach; semi-automatic tools; interrupted-flow and discrete-sampling platforms; single-enzyme research; multiple-enzyme research (M.E.A.); enzyme characterization; calculation of enzyme actions from software readings; generalized platforms for enzyme automation; Appendices I. released computerized enzyme assays, II. Terminology utilized in enzyme automation, III. equipment utilized in enzyme automation, IV. laptop application for generalized enzyme computerized procedure.

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Additional info for Automated Enzyme Assays

Example text

Enzyme is added to give a final concentration of 50 pg protein/ml, and after 10 min at 30 "C the fall in absorbance at 420 nm is measured. In detail, the actual assay proceeds in the following stages: 1. The stock solutions are prepared. (A stock solution is any solution that is used to prepare a reagent. ) 2. The reagents are prepared by dilution of the stock solutions. The convenient stock solutions in this case are 1 M potassium phosphate¶ 50 mM lactate and 10 mM ferricyanide. 75 mg protein/ml.

For example, changes in the heat content of the reaction mixture may be exploited, as in the calorimetric measurements of biological oxidations by Estabrook and Gutfreund (1960). Magnetic measurements may be used, for example in a study of myoglobin by Brill et al. (1960). There have been many studies on enzymes using NMR (nuclear magnetic resonance); Shulman (1962) summarized the use of electron and nuclear spin resonance in biology and gave some examples of its application to enzymology (see also Beinert and Palmer 1965).

As an example, Opher et al. (1966) studied the isoenzymes of serum lactate dehydrogenase by strip electrophoresis. They incubated the strips SEMI-AUTOMATIC METHODS 29 with substrate and tetra-nitro-blue tetrazolium. This was reduced to a purple formazan by the action of the enzyme, and the amount of reduction was proportional to enzyme concentration. Quantitative determination of the coloured area was carried out with a densitometer; the assay may therefore be regarded as a semi-automated procedure.

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