Isolation of bovine mammary lymphocytes for fluorescent activated flow cytometry.
A detailed methodology is described for the isolation of T lymphocytes from bovine mammary gland secretions for flow cytometry. Mammary gland secretions are collected and centrifuged to separate the leukocytes from the mammary supernatant. The leukocytes are purified using a density gradient and diluted for fluorescent staining. Using tri-color fluorescent staining, the T lymphocytes are identified by monoclonal antibodies and stained with secondary antibodies. Flow cytometry is used to quantify the cellular subpopulations of the T lymphocytes. Proportional and statistical analysis of the flow cytometric data is conducted with computer software, CELLQuest and SAS.
Ebling
T L
TL
Washington State University, Department of Animal Sciences, Pullman, WA, USA.
Fox
L K
LK
Byrne
K M
KM
D000818:Animals; D000911:Antibodies, Monoclonal; D001940:Breast / Q000166:cytology*; D002417:Cattle; D002469:Cell Separation; D002499:Centrifugation, Density Gradient; D005260:Female; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D013194:Staining and Labeling; D013601:T-Lymphocytes / Q000166:cytology*
D000911:Antibodies, Monoclonal