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Guanidine extracts of crude Bacillus anthracis cell wall were used to vaccinate BALB/c mice and to develop monoclonal antibody (MAb) to vegetative cell surface antigens. Two hybridomas selected during this study produced immunoglobulin M immunoglobulins, which appear to be directed to an epitope associated with the galactose-N-acetyl-D-glucosamine polysaccharide. Both demonstrated specificity in their binding to purified B. anthracis cell wall, o- stearoyl-polysaccharide conjugates, and intact, nonencapsulated vegetative cells. The interaction of the MAbs with purified polysaccharide was inhibited by 0.5 M galactose and lactose but not by N-acetylglucosamine, glutamate, glyciine, or glycerol. Inhibition by glucose or sucrose was approximately 75% of that seen with galactose. Electron microscopy showed that both MAbs interacted with the cell wall of vegetative cells as well as with the cortex of spores. Neither MAb reacted with encapsulated vegetative cells, such as those from infected guinea pigs, nor did they react with intact spores. After conjugation to fluorescein isothiocyanate, the MAbs stained intensely all B. anthracis strains tested, whereas with two exceptions, none of the strains of 20 other Bacillus spp. was stained. The exceptions, strains of Bacillus cereus, could be differentiated from B. anthracis by being beta-hemolytic on blood agar. Reprints. (kt), Availability: Pub. in Jnl. of Clinical Microbiology v28 n2 p223- 231, Feb 90. No copies furnished by DTIC/NTIS.

Sera from Hartley guinea pigs vaccinated with a veterinary live spore anthrax vaccine were compared with sera from guinea pigs vaccinated with the human anthrax, vaccine, which consists of aluminum hydroxide-adsorbed culture proteins of Bacillus anthracis V770-NP-1R. Sera from animals vaccinated with the spore vaccine recognized two major B. anthracis vegetative cell-associated proteins that were either not recognized or poorly recognized by sera from animals that received the human vaccine. These proteins, termed extractable antigens 1 (EA1) and 2 (EA2), have molecular masses of 91 and 62 kilodaltons, respectively. The EA1 protein appeared to be coded by chromosomal DNA, whereas the EA2 protein was only detected in strains that possessed the pX01 toxin plasmid. Both of the extractable antigen proteins were serologically distinct from the components of anthrax edema toxin and lethal toxin. Following vaccination with the live spore vaccine, the EA1 protein was the predominant antigen recognized, as determined by electrophoretic immunotransblots. Vaccine trials with partially purified EA1 demonstrated that it neither elicits protective antibody against anthrax nor delays time to death in guinea pigs challenged intramuscularly with virulent Ames strain spores. In addition, animals vaccinated with sterile gamma-irradiated cell walls had significant antibody titers to the N-acetylglucosamine-galactose polysaccharide of B. anthracis but were neither protected nor had a delay in time to death following challenge., Published in Infection and Immunity, v56 n2 p349-356, Feb 1988.

By using electrophoretic immuno-transblots, EITB (Western blots), we compared sera from Hartley guinea pigs vaccinated with a veterinary live-spore anthrax vaccine to those vaccinated with the human anthrax vaccine, consisting of aluminum hydroxide-absorbed culture proteins of Baccillus anthracis strain V770-NP-1R. Sera from animals vaccinated with the spore vaccine recognized two major B. anthracis vegetative-cell-associated proteins not recognized by sera from animals receiving the human vaccine. These proteins, termed extractable antigens 1 (EA1) and 2 (EA2), have molecular masses of 91 and 62 kilodaltons, respectively. The EA1 protein appeared to be coded by chromosomal DNA, whereas the EA2 protein appeared to be coded by chromosomal DNA, whereas the EA2 protein was only detected in strains possessing the pX01 toxin plasmid. Both of the EA proteins were serologically distinct from the three anthrax toxin components, as determined by monoclonal antibody to protective antigen, edema factor, and lethal factor, and specific antisera to the EA proteins.