Neutrophil Phagocytic Assay

Daniel Blount daniel_blount at hotmail.com
Tue Dec 18 18:36:06 EST 2001


Whilst looking through a message board in the past, I remember seeing an 
archive describing a neutrophil phagocytic assay. I know that there are kits 
on the market such as Phagotest(r), but I would like to have a go at an 
assay myself. The plan I have is to use fluorescein conjugated zymosan 
particles as the target and hence measure phagocytosis on the flow 
cytometer. I read that you can quench mebrane bound and free label with 
propidium iodide, however, I would prefer to have a control in which I add 
cytochalasin D, which inhibits phagocytosis. Does anybody have a protocol 
for a similar method? I need to know measurements such as the ratio of 
neutrophils to zymosan, and the amount and time of incubation for 
cytochalasin D. Do I need to attach complement to my zymosan? How is best to 
do this? (time and temperature of incuabtion etc.) Also, would it be 
possible to carry this assay out on whole blood (maybe with a RBC lysis 
step)?

Many thanks

Daniel G. Blount
Nottingham University
School of Pharamceutical Sciences
University Park
Nottingham
NG7 2RD

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