Abstract: A mutant strain of E. coli (44110-1) was used in the cup-plate method for ultra-micro analysis of vitamin B₁₂. This method has been proved to be sensitive while the concentration of B₁₂ is within the range of 0.05—0.5γ/ml. B₁₂ contents in several samples of commercial liquid extract of liver were thus assayed and found to be approximately the same to that labelled. The cup-plate method of employing E. coli as an assay organism is superior to that using Lactobacilli, for culture media and instruments used are easily obtained and the whole assay is not difficult to control.