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Serum vitamin B12 levels were measured in 300 Indian subjects, comprising 165 haematologically normal volunteers, 78 cases of nutritional megaloblastic anaemia and 57 patients with myeloproliferative disorders. Each sample was assayed by three different techniques: i) a microbiological assay using Euglena gracilis as the test organism; ii) radioisotope dilution assay, using pure intrinsic factor as the B12 binder and haemoglobin-coated charcoal for separation of bound from free vitamin B12; iii) radioisotope dilution assay using intrinsic factor as the vitamin B12 binder and DEAE cellulose for separation of bound from free vitamin B12. Results by the three methods correlated closely at normal and high levels of serum B12: at low levels microbiological assay and radio-assay using haemoglobin-coated charcoal correlated well but DEAE cellulose failed to detect 16 of 68 patients with vitamin B12 deficiency. The radioisotope dilution assay tended to yield higher values of serum vitamin B12 than did the microbiological assay. In none of the healthy volunteers was a clearly low value found by any of the techniques, but 20 of them had low serum B12 levels. Radioisotope dilution assay using haemoglobin-coated charcoal is recommended as an accurate assay procedure for developing countries like India, where ready-made kits are not easily available and vitamin B12 deficiency states are common.