A. Johannisson et al., FLOW-CYTOMETRIC STUDIES OF THE PHAGOCYTIC CAPACITIES OF EQUINE NEUTROPHILS, Acta veterinaria Scandinavica, 36(4), 1995, pp. 553-562
Methodological aspects of flow-cytometric evaluation of the phagocytic
properties of equine neutrophils were elucidated. The kinetics of att
achment and ingestion were studied, and the phagocytic process was mor
e rapidly completed when serum-opsonized yeast cells were used than wi
th use of IgG-opsonized yeast cells. Trypan blue was successfully used
to quench fluorescence of non-ingested yeast cells. There were only m
inor differences in the kinetics of phagocytosis between quenched and
unquenched samples, indicating that attachment is rapidly followed by
ingestion. Trypan blue quenching caused loss of cells with light scatt
ering properties of granulocytes, although this did not affect the det
ermined frequencies of truly phagocytic neutrophils. Aggregation of ye
ast cells proved to be a disturbance but not an obstacle to the determ
ination of frequencies of actively phagocytic cells. Flow cytometry is
well suited for studies of phagocytosis of yeast cells by equine neut
rophils, and the trypan blue quenching provides a means of eliminating
false-positive events due to aggregation of yeast cells. The main adv
antage of the flow-cytometric method is the possibility of rapid proce
ssing of a large number of samples, making the method useful for studi
es of herds.