Flow cytometric differentiation of avian leukocytes and analysis of their intracellular cytokine expression.
A flow cytometric method for the identification of chicken blood leukocyte subpopulations and thrombocytes was developed. An anti-chicken CD45 phycoerythrin-labelled antibody was used to separate leukocytes from red blood cell nuclei. Leukocytes and thrombocytes were identified using a combination of their CD45-positivity and their typical side scatter properties. The identity of the CD45-positive cells was confirmed by sorting the subpopulations and subsequent light microscopic evaluation. In these differentiated cell populations, intracellular expression analysis of the proinflammatory cytokines interleukin-1beta and interleukin-6 was subsequently optimized on whole blood after in vitro stimulation with lipopolysaccharide from Escherichia coli strain O127:B8.
De Boever
S
S
Department of Pharmacology, Toxicology and Biochemistry, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium. sandra.deboever@ugent.be
Croubels
S
S
Demeyere
K
K
Lambrecht
B
B
De Backer
P
P
Meyer
E
E
D000818:Animals; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism; D002469:Cell Separation; D002645:Chickens; D004912:Erythrocytes / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods*; D053583:Interleukin-1beta / Q000378:metabolism*; D015850:Interleukin-6 / Q000378:metabolism*; D017493:Leukocyte Common Antigens / Q000276:immunology / Q000378:metabolism*; D007962:Leukocytes / Q000166:cytology / Q000378:metabolism; D010799:Phycoerythrin; D013194:Staining and Labeling
D000911:Antibodies, Monoclonal; D053583:Interleukin-1beta; D015850:Interleukin-6; D010799:Phycoerythrin; D017493:Leukocyte Common Antigens