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An investigator here is comparing the effects of activated [using phorbol]
vs non-activated human PMN, but whereas they expected (based on other
studies) the activated PMN to have an enhancing effect compared to the
unactivated PMN, they find similar results from the addition of either
population.

Their PMN isolation procedure involves dilution of blood 1:2 with saline,
addition of dextrin to 5%, allowing cells to settle for 45 min at 37 C,
taking the supernate and underlaying it with Histopaque, and centrifuging
for 30 min (I don't know the rpm).  The PMN are recovered as the pellet.  [I
believe I've described this accurately]

The markers they are looking at, on human red blood cells, are percentages
of RBC expressing CD36, CD49, and phosphatidyl serine.  The activated PMN
were expected to increase these values compared to unactivated, i.e., non
PMA treated, PMN, when incubated with the RBC.

Would the isolation procedure described above be expected to activate PMN
without the addition of PMA?

If so, does anyone know of a gentler isolation procedure which might
not/does not activate PMN?

Thanks for any thoughts.

Ray

Ray Hester
Univ. of South Alabama
Mobile, AL 36688
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