Purified lipopolysaccharide (LPS) extracted with phenol-water from smooth Brucella abortus was hydrolyzed with 1% acetic acid at 1000C. The degraded polysaccharide (AH) released gave reactions of identity with the native polysaccharide hapten (NH) in phenol-wateror trichloroacetic acid-extracted endotoxin preparations of B. abortus and with the polysaccharide (poly B) extracted by trichloroacetic acid from rough B. melitensis strain B115. Poly B was present in the soluble cytoplasmic fraction but not in the membrane fraction, of disrupted B115 cells. It could not be extracted from three rough mutants of B. abortus or from B. canis or B. ovis cells. Both AH and NH shared determinants present on smooth LPS and missing from poly B. Sugars found in purified LPS, NH, and AH included mannose, glucose, quinovosamine, glucosamine, and 2-keto-3-deoxyoctonate. Poly B contained only a trace amount of quinovosamine and no 2keto-3-deoxyoctonate detectable by the thiobarbiturate assay. Sera from some rabbits immunized with pure smooth LPS and some, but not all, cows infected with field strains of B. abortus recognized the determinants missing from poly B. A subclass-specific enzyme-linked immunoassay showed that most of the antibody in sera from infected cows which binds to smooth LPS and to NH is of the immunoglobulin Gl subclass.

The smooth lipopolysaccharide complex of the outer surface of smooth Brucella abortus cells is believed to be the antigenic component involved in serological tests routinely used for the diagnosis of brucellosis. Sera from cattle vaccinated or infected with B. abortus generally contain antibody directed toward the smooth lipopolysaccharide complex. The brucella organism contains a large number of other antigenically distinct components. The biological significance of some of these antigens has been demonstrated by showing that sera from infected cattle have precipitins to these components. These sera revealed up to seven distinct lines in immunoelectrophoresis with a protein-rich antigen mixture prepared from rough strain B. abortus 45/20, whereas sera from strain 19-vaccinated cattle did not reveal these lines at 4 or more months after vaccination. Monospecific antisera were prepared against six antigens in this mixture, and the purification of two of them by antibody affinity chromatography is described. tion of cells with hypertonic NaCi as described by Jones et al. (14) or by sonic disruption as described by Hinsdill and Berman (13).