Isolation of bovine neutrophils with biomagnetic beads: comparison with standard Percoll density gradient isolation methods.
A prerequisite for studies on bovine neutrophils is a reliable method of neutrophil isolation from blood to obtain highly purified cell populations that are functionally active. Since current techniques of neutrophil isolation fall short of these requirements, we have developed a newer and more effective technique for isolation of bovine neutrophils that utilizes biomagnetic beads coated with a monoclonal antibody that recognizes an abundant surface antigen on bovine neutrophils to purify these cells. Comparison of the purity and viability of bovine neutrophils isolated by a conventional method (continuous Percoll density gradient) with this new method showed that neutrophils isolated with biomagnetic beads were higher in purity and had an increased yield. In addition, cells isolated with biomagnetic beads demonstrated normal or even improved function in assays of chemotaxis, phagocytosis, degranulation, and respiratory burst activity. Finally, bovine neutrophils isolated using this method showed an overall lower level of spontaneous apoptosis, which correlates well with the high level of viability observed in the purified cell preparations. Thus, this method represents a significant advance over current methods for isolating bovine neutrophils and would be widely applicable to labs studying the biochemistry and signal transduction pathways in these cells.
Soltys
J
J
Department of Veterinary Molecular Biology, Montana State University, Bozeman 59717, USA.
Swain
S D
SD
Sipes
K M
KM
Nelson
L K
LK
Hanson
A J
AJ
Kantele
J M
JM
Jutila
M A
MA
Quinn
M T
MT
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D017209:Apoptosis; D018821:CD18 Antigens / Q000096:biosynthesis; D002417:Cattle; D015550:Cell Degranulation; D002499:Centrifugation, Density Gradient / Q000379:methods*; D002633:Chemotaxis; D018189:Immunomagnetic Separation / Q000379:methods*; D019041:L-Selectin / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D009504:Neutrophils / Q000166:cytology / Q000276:immunology / Q000378:metabolism / Q000502:physiology; D010587:Phagocytosis; D011205:Povidone; D016897:Respiratory Burst; D012822:Silicon Dioxide
D000911:Antibodies, Monoclonal; D018821:CD18 Antigens; D019041:L-Selectin; C016039:Percoll; D012822:Silicon Dioxide; D011205:Povidone