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  • Author or Editor: Thomas J. Inzana x
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Summary

Immunogenicity of the lipid A component of Haemophilus somnus lipooligosaccharide in cattle and mice was examined after purification, detoxification, and covalent conjugation to a protein carrier. After 2 inoculations, a substantial antibody response was induced in most cattle to lipid A and the protein carrier. To determine whether antibodies to lipid A would be protective, 5 x 107 colonyforming units of H somnus strain 649 were administered IV to endotoxin-responsive (C3H/HeN) mice. In one study, 8 of 13 C3H/HeN mice aborted when inoculated. In contrast, abortion did not result when mice were inoculated with the same dose of an isolate of H somnus normally found in the prepuce or with the rough mutant Escherichia coli J5. In addition, endotoxin-nonresponsive (C3H/ HeJ) mice were significantly (P =0.03) more resistant to abortion by strain 649 than were C3H/HeN mice, but inoculated C3H/HeN mice were only slightly more resistant to H somnus abortion, compared with control mice. Although a large antibody response to lipid A was detected, there was no significant difference in the immunized group between mice that aborted and mice that delivered normally. Thus, lipooligosaccharide and other properties of virulent H somnus strains may contribute to abortion in mice.

Free access
in American Journal of Veterinary Research

Summary

An efficient, single-step method for purification of the 110-kilodalton (kDa) hemolysin of Actmobacillus pleuropneumoniae was developed. An immunoaffinity column was made by cross-linking murine monoclonal antibody 8C2 to the 110-kDa hemolysin of A pleuropneumoniae strain J45 serotype 5 to protein A-agarose beads. Purified hemolysin with high hemolytic activity was obtained after washing the column with phosphate-buffered saline solution, and eluting the hemolysin with 50 mM diethylamine, pH 11.0. The same column was also used to purify the hemolysin from A pleuropneumoniae strain 4074 serotype 1. The purification procedure could be completed within 5 hours, and almost 50% of the total hemolytic activity and hemolysin protein was recovered in pure form.

Free access
in American Journal of Veterinary Research

Abstract

Objective—To characterize matrix metalloproteinase (MMP)-2 and -9 in CSF of clinically normal dogs.

Sample Population—Samples of CSF collected from 23 dogs.

Procedure—Dogs were anesthetized, CSF samples were collected, and dogs were then euthanatized. Each CSF sample was evaluated immediately for RBC count, WBC count, and protein and glucose concentrations, and cytologic examination also was performed. Samples were considered normal when protein concentration was < 25 mg/dL and CSF contained < 6 WBCs/μL and < 25 RBCs/μL. Samples were stored at –70°C. Sections of brain tissue were collected and processed for histologic examination. The MMPs were evaluated by use of gelatin zymography and a polyclonal antibody-based sandwich ELISA.

Results—Mean WBC count for CSF samples was < 1 WBC/μL (range, 0 to 3 WBCs/mL). Mean protein concentration was 12 mg/dL (range, 8 to 17 mg/dL). Mean RBC count was 3.65 RBCs/μL (range, 0 to 21 RBCs/μL). All CSF samples generated a clear band on zymography gels that corresponded to the human commercial standard of proenzyme MMP-2. Other major clear bands were not detected on zymography gels. Bands correlating to MMP-9 were not detected in any samples. The ELISA results revealed a mean ± SD proenzyme MMP-2 concentration of 5.61 ± 1.92 ng/mL (range, 3.36 to 10.83 ng/mL).

Conclusions and Clinical Relevance—The proenzyme form of MMP-2 is detectable in CSF of clinically normal dogs, whereas MMP-9 is not detectable. Additional investigation of MMPs in CSF from dogs with various diseases of the nervous system is indicated. (Am J Vet Res 2002;63:1359–1362)

Full access
in American Journal of Veterinary Research