Tracer method vitamin

The low-molecular-weight vitamin biotin is easily conjugated to antibodies and enzyme markers. Up to 150 biotin molecules can be attached to one antibody molecule, and the strong affinity of the biotin for the glycoprotein avidin allows its use as complex-ing secondary reagents. Biotin labeling of the primary (direct) or secondary (indirect) antibody can be used in the avidin-biotin methods. In the labeled avidin method the tracer is attached directly to the avidin molecule. In the avidin-biotin bridge method a biotinylated enzyme such as peroxidase is allowed to bind after attachment of avidin to the biotin-labeled antibody. 

Variability in Absorption Estimates In this study, the occurrence of a negative absorption value for one subject and the absence of a significant vitamin C effect raise some questions about the accuracy of the method However, the expected changes in absorption due to dietary treatments may be masked by the analytical variations associated with absorption measurements and biological variabilities of iron absorption Analytical variations can be introduced at several stages of the analytical procedures incomplete fecal collection, inhomogeneous samples, iron contamination, incomplete colorimetric reaction, non-quantitative recovery after chemical ashing, and variations in isotopic measurements due to ion statistics, memory effects, instrument drift, etc Some of these are not as serious as others, for example, contamination with natural iron woiold not affect the estimate of tracer concentrations provided it occurs before the total iron content is measured ... 

Immunoassays, specific protein-binding assays, and radioisotope tests are sometimes used for the determination of water-soluble vitamins. These are the only feasible and practical methods for the quantification of certain vitamins in physiological samples. The principle of competitive protein binding (CPB) using labeled radioactive or fluorescent tracer is still routinely applied to quantification of serum vitamin... 

The vitamin B12 activity of a given sample may be determined by biological, chemical, microbiological or physical methods including radioactive tracer, spectrophotometric, and partition techniques. The nature of the sample, the concentration of activity, precision, and time required will usually determine the assaj of choice. 

A radioisotope dilution assay and an assay based on animal growth are the most reliable methods for the determination of vitamin B12 activity. The tracer technique is highly specific for cyanocobalamin or analogs which are convertible to cyanocobalamin. The assay is specific for cyanocobalamin if cyanide treatment of the sample is avoided total cobalamins convertible to cyanocobalamin are determined if a given sample is first treated with cyanide. The method consists in the addition of a known amount of pure [ Co]-cyanocobalamin. A series of selective extractions and adsorptions to remove interfering substances is completed and the radioactivity and color of the purified sample are measured. From these data, it is possible to calculate the amount of cobalamin present in the original sample. The isotope dilution method is accurate and precise and can be used for both relatively pure samples and for crude extracts of low potency. 

The quantitation of riboflavin together with thiamine and niacin using HPTLC silica gel plates and methanol/water (70 30, v/v) as mobile phase was described by Diaz et al. (30). For riboflavin the native fluorescence was used and a preplate derivatization was applied for the other two vitamins (addition of a fluorescent tracer to label nicotinic acid conversion of thiamine into thio-chrome). The developed plates were scanned by a commercially available bifurcated flber-optic-based instrument that transferred the excitation and emission energies between the plate and the fluorescence spectrometer. Calibration curves for the determination of riboflavin 48 to 320 ng, thiamine 300 to 750 ng, and niacin 10 to 100 ng were established. The advantages of this method are that no elimination of excess oxidation reagent is necessary and that the simultaneous determination of vitamins with only one detector is possible. 

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