was determined in biological materials by three basically different methods: microbiological assa) with Luctobaeiltus lcichmcmnii, microbiological assay with Esehcrichiu coli and radioassay. The method with E. coli has a relatively low sensitivity to vitamin B,, and in some cases of vitamin B,, determination in microbial materials it can be used only after a separation of the interfering suhstances by gel chromatography. The procedure is suitable for orientational determinations of vitamin B,2 because it is very little affected by external factors. 'The assay with L. Irich/nnnnii is universal owing to its high specifity and sensitivity to vitamin B,2. The main disadvantage of the latter procedure depends on the high requirements for a clean atmosphere which can be mained in laboratories in industrial areas only with difficulties. These limitations do not apply to the quick and sensitive radioassay. The radioassay can be used after a suitable adjustment of the working procedure for large series of analyzes of biological materials without any preliminary separational techniques. The test can be carried out in normal laboratories providing that the principles of work with radionuclides are obeyed.Man) physicochemical. chemical and microbiological methods are described for vitamin B12 analysis. The determination of this vitamin in biological materials is complicated in many cases owing to its low concentration in the medium and to the presence of interfering substances. The majority of chemical and physicochemical procedures can be applied only for pharmaceutical preparations or for various concentrates and only if the interfering substances were preliminarily removed [ 1,2]. The microbiological assays based on the growth of a test microorganism, that is dependent on vitamin B,,. and that can without any separational pretreatment be applied for vitamin B,, determination even in media with extremely low vitamin B,, levels, i.e. in biological materials. Although the microbiological assay is highly specific, the possibility that the test microorganism might utilize some components of the biological material asgrowth factors must always be taken into account [3--61.Clinical biochemistry has therefore recently often been using methods for vitamin B,, which join the high specifity of microbiological assays with the high sensitivity of radiometric methods, that is the competitive protein binding analysis (CPBA) [7,8] and the radioimmunoassay (RIA) 191. The analysis of vitamin B,, by these two methods lasts only a few hours. The methods are based on the competition between vitamin B,, labelled with a radionucleotide and between normal viGmin B,, to be bound to the binding protein (CPBA) or to an antigen (RIA). The CPBA method is quicker than the microbiological assays. Neither antibiotics nor other substances affecting the microbial growth interfere with the CPBA procedure. The method can easily be standardized and is highly suitable for routine analysis. It has mainly been used [lo-141 in clinical biochemistry but very seld...
