Malleo-proteins from synthetic broth mallein of six strains of Pseudomonas mallei were separated by trichloroacetic acid precipitation, amonium sulfate precipitation and Ultrogel AcA 34 gel filtration chromatography. When tested comparatively with Dutch PPD mallein as standard on P. mallei-sensitized and normal horses all the strains were found to be malleinogenic, trichloroactive acid precipitated proteins were comparable to Dutch PPD mallein in potency and innocuity whereas ammonium sulfate-precipitated proteins elicited non-specific reactions. Ultrogel AcA 34 chromatographed high molecular weight proteins (MW range > 350 000) were having equal or higher potency and without adverse reactions and low molecular weight proteins (MW range 120 000-170 000) nearly inactive. Ultrogel AcA 34 column fractionation studeis revealed that mallein activity was associated with high molecular weight proteins and represented a purer sensitin than PPD mallein for the purpose of mallein test. {\textcopyright} 1994.