The reactivities of the two cysteine thiol groups of calf thymus F3 histone were investigated using 5,5′-dithiobis-[2-nitrobenzoic acid], (DTNB). In isolated histone, both thiol groups were available for reaction. However, analysis of reaction profiles of native deoxyribonucleohistone, (DNH), in various solvent conditions, together with gel electrophoresis studies of DNH modified with DTNB, showed that only one of the thiol groups is normally modified by the reagent. If NaCl is present (above 1.0M) the other thiol group can also be modified. The reactivities of both groups were largely independent of the degree of DNH supercoiling and of the binding of F3 to the DNA. © 1974 Oxford University Press.