R. K. Anderson

BRUCELLA-AGGLUTINATING ANTIBODIES: RELATION OF MERCAPTOETHANOL STABILITY TO COMPLEMENT FIXATION.

Brucella-agglutinating antibodies from selected bovine blood serums and milk samples did not fix complement, and after treatment with 2-mercaptoethanol they lost agglutinating power. After infection of calves with Brucella abortus, strain 19, agglutinins for Brucella that were inactivated by mercaptoethanol appeared earlier than those stable to mercaptoethanol. Under the conditions of these experiments, the appearance and development of complement-fixing capacity coincided closely with the mercaptoethanol stability of the agglutinins for Brucella.

Application of a whole-blood lymphocyte stimulation test in detection of Brucella infection in an outbreak

A whole-blood lymphocyte stimulation test (WBLST), the standard plate and tube agglutination tests, the Brucella buffered antigen test, the 2-mercaptoethanol agglutination test, the Rivanol plate precipitation test and bacteriological isolation were utilized in a brucellosis outbreak investigation in a beef herd. Three of the animals were classified as not infected serologically. However, these 3 animals were classified as infected on the WBLST, andBrucella abortus biotype 1 (not strain 19) was isolated from their lymph nodes. The WBLST exhibited significant sensitivity in this investigation and more observations of this nature might strengthen the application of this assay in similar situations.

In vitro stimulation of bovine milk lymphocytes standardization of the assay for bovine brucellosis

Abstract Lymphocytes of bovine milk origin were investigated by immunostimulation in vitro to standardize the assay for measuring the immune responses of the cells which might be useful in further understanding the immunopathology and diagnosis of bovine brucellosis. The lymphocytes were separated from whole freshly collected milk by centrifugation. The pellet of lymphocytes was washed in RPMI-1640 medium, cultured at different concentrations for different days and with Brucella abortus soluble antigen strain 1119-3 and Concanavalin A. Each culture was labelled with 1.0 μCi of methyl-[3H]thymidine 16–18 hours prior to termination of incubation at 37 C. Termination was done by cooling to 4 C. The cells were harvested for liquid scintillation counting spectrometry. In the groups of calfhood vaccinated cows and nonexposed milkers, a milk lymphocyte concentration of 2.0 × 106/ml of medium yielded a statistically significant blastogenesis. The Brucella abortus soluble antigen concentration of 4.4 μg of protein/well was found optimal to induce significant immunostimulation. A period of 4 days of incubation of the milk lymphocyte in the test was found optimal in inducing statistically significant blastogenesis in this system.

Specific in-vitro lymphocyte immunostimulation activities of Brucella abortus fractions obtained by column chromatography

Abstract A Brucella abortus soluble antigen (BASA) preparation and fractions obtained thereof, by column chromatography, were compared in terms of their ability to induce specific lymphocyte stimulation responses (LSR) in lymphocytes from cattle infected with B. abortus . Endotoxin and protein contents and the fractions were determined. The LSR induced by BASA and the fractions were compared in terms of correctly identifying samples from infected and non-infected cattle. The sensitivity and specificity for each preparation were determined and these two attributes were then correlated with endotoxin and protein content. The results suggest that lymphocyte stimulation had greater association with relative protein content than endotoxin content of the antigen preparations.

Application of indomethacin as a potentiator of lymphocyte blastogenesis in Brucella abortus exposed cattle

Lymphocytes from Brucella abortus field strain infected, strain 19 vaccinated, non-exposed and field strain infected, but immunologically unresponsive cattle were incubated with B. abortus antigen and indomethacin. There were significant increases (P less than 0.005) in the blastogenic responses, as measured by [3H] thymidine uptake, in cultures with indomethacin as compared to cultures without indomethacin. Lymphocyte blastogenic responses to B. abortus antigen were potentiated by indomethacin in both B. abortus exposed and non-exposed cultures. However, potentiation of sensitized lymphocyte blastogenic responses by indomethacin was significantly greater (P less than 0.005) than that in non-exposed lymphocytes. Additionally, indomethacin significantly potentiated Brucella-induced lymphocyte blastogenic responses in lymphocytes from anergic cattle.

Long-term storage of blood prior to whole-blood lymphocyte culturing

Abstract Studies were conducted to investigate the effect on blastogenesis of long-term storage of blood prior to whole-blood lymphocyte culturing. Peripheral blood from normal cattle was utilized. Blood from each animal was divided into 2 parts, A and B, following collection. Part A was left intact while RPMI-1640 culture medium was added to part B immediately following collection. Both parts were then kept at room temperature for a total of 7 days and a portion of each blood was tested every day. The cultures were incubated and assayed for [3H] thymidine incorporation into their DNA. It was observed that intact blood (A) was good for 4 days but deteriorated thereafter. Addition of RPMI-1640 to B prolonged the keeping quality of blood for 7 days prior to culturing. The possible application of these findings are discussed.

The optimal time to stain for noncytopathic bovine virus diarrhea field sample virus using indirect fluorescent antibody technique.

Abstract The use of an indirect fluorescent antibody (IFA) technique has been further explored to determine the optimum day(s) to stain for a noncytopathic bovine virus diarrhea virus from pooled and individual field samples. The IFA test further confirmed that under the experimental conditions of this study, somewhere between days 4 and 5 would be an optimum time to stain for a noncytopathic BVD virus which was present in tissues of suspected animals and diluted with minimal essential media.