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Comparison of soluble antigens Leptospira interrogans serovars by SDS-PAGE, crossed immunoelectrophoresis and immunoblotting.
1992
Baik Y.O. | Mah J.S.
Rapid purification of a 110-kilodalton hemolysin of Actinobacillus pleuropneumoniae by monoclonal antibody-affinity chromatography
1992
Ma, J. | Inzana, T.J.
An efficient, single-step method for purification of the 110-kilodalton (kDa) hemolysin of Actinobacillus 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.
Mostrar más [+] Menos [-]Evaluation of total protein content in tears of dogs by polyacrylamide gel disk electrophoresis
1992
Barrera, R. | Jimenez, A. | Lopez, R. | Mane, M.C. | Rodriguez, J.F. | Molleda, J.M.
Concentration of total proteins was measured and sodium dodecyl sulfate-polyacrylamide gel disk electrophoresis was performed on tear and plasma samples obtained from 26 healthy dogs, and the results were compared. Mean +/- SEM concentration of total proteins in tears was 0.63 +/- 0.04 g/dl, and significant effects of age or gender were not found. The protein composition of tears in dogs was complex, and bands from light and heavy chains of immunoglobulins were identified by electrophoresis.
Mostrar más [+] Menos [-]Characterization of a feline T-cell-specific monoclonal antibody reactive with a CD5-like molecule
1992
Ackley, C.D. | Cooper, M.D.
The 43 monoclonal antibody raised against feline T cells was found to react with a single-chain glycoprotein of Mr 72,000 that is present on most thymocytes, 60% of lymph node cells, 20% of splenocytes, and 45% of blood mononuclear cells. All CD4+ and CD8+ T cells were found to express the 43-reactive determinant, as did a small subpopulation of CD4-/CD8-/IgM- lymphocytes in the periphery. The 43-reactive determinant was not detected on B cells, macrophages, or other types of blood cells. The 43 antigen was phosphorylated in resting and activated T cells. Its expression was upregulated by stimulation with phorbol myristate acetate and with phytohemagglutinin. When added to concanavalin A-stimulated T-cell cultures in low concentrations, the 43 antibody was found to augment mitogenesis. The data indicate that this antibody may identify a CD5 homologue on feline T cells.
Mostrar más [+] Menos [-]Polypeptides associated with Pasteurella multocida infection in rabbits
1992
Zimmerman, T.E. | Deeb, B.J. | DiGiacomo, R.F.
Polypeptides from whole cell preparations of Pasteurella multocida serotypes A:12 and A:3 were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose paper. Antigens were detected by immunoblot analysis, using sera from 3 groups of rabbits. Sera were obtained from rabbits inoculated intranasally with P multocida serotype A:12 or A:3, from rabbits maintained in a rabbitry with enzootic P multocida A:12 infection, and from rabbits maintained in a rabbitry with enzootic P multocida A:3 infection. Immunoblot analyses of pre- and postinoculation sera from experimentally infected rabbits, using serotype A:12 antigen, revealed 3 polypeptides with approximate molecular mass of 28, 30, and 37 kDa that consistently detected antibodies after P multocida-induced infection. Sera from rabbits naturally infected with either serotype, tested against serotype A:12 and A:3 antigens, detected the same polypeptides in both serotypes. Thus, immunologic reactivity to these polypeptides may be useful for serologic detection of P multocida infection.
Mostrar más [+] Menos [-]Determination of protein concentrations and their molecular weight in tears from cats with normal corneas and cats with corneal sequestrum
1992
Davidson, H.J. | Gerlach, J.A. | Bull, R.W.
Protein concentration was determined, using the Bradford technique, in tears from cats with normal corneas and from cats with corneal sequestrum. Tears from the former group contained 5.81 +/- 2.29 mg of protein/ml; those from cornmeal sequestrum-affected cats contained 6.21 +/- 2.21 mg/ml. Difference between the 2 values was not significant. Molecular weight determination was made, using 4 to 20% sodium dodecyl sulfate-polyacrylamide gels. Molecular mass of proteins ranged from 263 to 14 kDa. There was no detectable difference in the band patterns for the 2 groups.
Mostrar más [+] Menos [-]Double-antibody sandwich enzyme-linked immunosorbent assay and immunoblot analysis used for control of caseous lymphadenitis in goats and sheep
1992
Laak, E.A. ter | Bosch, J. | Bijl, G.C. | Schreuder, B.E.C.
A double-antibody sandwich ELISA for detection of antibodies directed against the exotoxin of Corynebacterium pseudotuberculosis, the cause of caseous lymphadenitis (CL) in small ruminants, was developed. A concentrated exotoxin was used. For interpretation of ELISA results, these sera were tested: sequentially obtained sera of C pseudotuberculosis-inoculated goats and sheep that were monitored for 68 weeks; sequentially obtained sera from 80 goats of 3 flocks with CL; sera from 652 goats of 7 flocks without CL; sera from 160 sheep of 4 flocks without CL; and 2,265 caprine and 208 ovine sera submitted for diagnostic testing. Data regarding the infection status and history of 10,454 of the 23,302 animals were collected after testing; most of these were goats that had been part of a CL control program. Specificity and sensitivity of the ELISA were nearly 100%. Subsequently, 31,978 animals from which no data on infection status of flocks had been collected were then tested. It was concluded that the ELISA is a useful diagnostic test for CL eradication programs. Sera with doubtful or inconclusive ELISA results were examined by use of immunoblot analysis. Proteins from C pseudotuberculosis culture supernatant were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted onto nitrocellulose. Six proteins with molecular mass of 68, 65, 39, 38, 31, and 29 kDa reacted with sera from goats and sheep with experimentally induced or naturally acquired infection. Immunoblot analysis was valuable in classifying sera with doubtful or inconclusive results by ELISA.
Mostrar más [+] Menos [-]Purification of the ninth component of the bovine complement cascade
1992
Eisenschenk, F.C. | Houle, J.J. | Hoffmann, E.M.
Conditions for purification of the ninth component of bovine complement (C9) were established. The conditions for binding and elution from diethylaminoethyl cellulose and hydroxylapatite were different than for human C9. Serum albumin, a frequent contaminant of bovine C9 preparations, was removed by chromatography on reactive-red agarose. The calculated molecular weight of bovine C9 was 66,000, and reduction with 2-mercaptoethanol affected its migration on polyacrylamide gel electrophoresis. Some preparations of bovine C9 migrated as 2 bands when partially reduced, but extensively reduced preparations had a single band.
Mostrar más [+] Menos [-]Prevalence of autoantibodies to thyroglobulin, thyroxine, or triiodothyronine and relationship of autoantibodies and serum concentrations of iodothyronines in dogs
1992
Thacker, E.L. | Refsal, K.R. | Bull, R.W.
Assays were developed to detect and measure antibodies (AA) to thyroglobulin (Tg) and to the thyroid hormones, thyroxine (T4) and triiodothyronine (T3). An ELISA to detect AA to Tg was developed, using purified canine Tg as the antigen and goat anti-canine IgG conjugated with alkaline phosphatase as the second antibody. A highly charged agarose electrophoresis assay was used for determination of AA to T4 and T3. Sera from dogs (n = 119) with clinical signs consistent with hypothyroidism were tested for AA to Tg, T4, and T3. Autoantibodies to at least 1 of the 3 thyroid antigens were detected in 58 of the 119 (48.7%) sera tested. Autoantibodies to Tg were detected more frequently in samples with low serum concentrations of thyroid hormones than in samples with normal concentrations. The presence of AA to T4, T3, or both was not significantly associated with low thyroid hormone concentrations, but this lack of association may have been attributable to binding of AA in the measurement of thyroid hormones by radioimmunoassay.
Mostrar más [+] Menos [-]Comparison of antibody responses in cattle to outer membrane proteins from Pasteurella haemolytica serotype 1 and from eight untypeable strains
1992
Simons, K.R. | Morton, R.J. | Fulton, R.W. | Confer, A.W.
Membrane associated proteins from 8 untypeable Pasteurella haemolytica strains were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and compared with those of P haemolytica serotypes 1 and 2. Cattle antisera obtained from P haemolytica serotype 1 vaccine trials were used in immunoblotting assays to compare the membrane proteins from the 8 untypeable strains with those from P haemolytica serotypes 1 and 2. Densitometry was used to identify bands, and using linear regression analyses, the peak area optical densities (measuring antibody response) were correlated to lesion scores from the vaccinated calves. Significant antibody responses to proteins of 99, 69, 60, 55, 47, 45, 39, 33, 30, 16, and 14.5 kDa were detected for 4 or more of the 8 P haemolytica untypeable strains. Serotypes 1 and 2 of P haemolytica contained a comigrating 30-kDa protein. Antibody responses to proteins of 39, 33, and 32.5 kDa were significant for 3 of the untypeable strains and had significant correlation to lesion scores. Antibody responses to various other proteins were significant for 2 untypeable strains each.
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