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Radioisotope dilution assay methods have attracted much attention since the first analytical symposium on this subject ten years ago. Isotope dilution assays of a wide variety of substances, including many biologically important substances such as vitamins, antibiotics, and insecticides, have been developed. An isotope dilution assay for vitamin B12 is included in the first U. S. Pharmacopeia XV supplement. Conditions which must be satisfied if dilution assays are to be reliable are enumerated. Tracer radiation stability and absence of exchange reaction must be assured. Several variants of the isotope dilution procedure have been proposed: the double (or successive) dilution and the reverse isotope dilution methods. The derivative method, which makes use of a radioactive reagent of known specific activity, can be applied directly if quantitative isolation procedures are known, or in a “derivative dilution” or reverse isotope dilution procedure. Reverse isotope dilution procedures, commonly used in biological investigations, are a convenient means for determining the purity of a labeled compound or its stability in complex media. The stability of vitamin B12 in dry cereals at concentrations as low as 0.46 γ per ounce has been demonstrated by this method.
Merck Sharp & Dohme Research Laboratories, Div. of Merck & Co., Inc., Rahway, N. J.