The cellular fatty acids of reference strains of most of the currently recognized species of Neisseria and Branhamella were examined. Analysis of fatty acids with chain lengths of over 12 carbons supported the division of the species into 2 groups. Group I, comprised of N. meningitidis, N. gonorrhoeae, N. flava, N. subflava, N. perflava, N. sicca, N. mucosa, N. lactamica and N. cinerea, contains methyl laurate, methyl palmitoleate, methyl palmitate, methyl oleate and, often, methyl myristate as principal fatty acids. Group II, represented by N. caviae, N. cuniculi, N. ovis and B. catarrhalis, contains large amounts of a 17-carbon fatty acids with a retention time similar to those of methyl-14, methyl hexadecanoate, methyl heptadecanoate and, often, methyl stearate in addition to those fatty acids found in group I organisms. The greatest distinction between the 2 groups was the percentage of major fatty acids with chain lengths > 16 carbons. Aminopeptidase activity was most useful in differentiating N. meningitidis from N. gonorrhoeae, N. cuniculi, N. ovis, N. caviae and B. catarrhalis had similar aminopeptidase reactions. Pigment profiles were of limited taxonomic value but were useful in differentiating between selected pigmented species.