 |
 |
 |
 |
There are many pitfalls in doing this type of experiment.
Obviously, if you are stimulating with an antibody and cross-linking with
another antibody you must make sure there is positivly no cross reaction with
the cross linking antibody and your phenotyping antibodies, a tiny bit of
cross-reactivity could have little effect with one antibody but you may get a
synergistic effect with all three.
Also the phenotyping antibodies may have a calcium mobilisation effect either
alone or synergistically.
Perhaps with enough controls you may be able to produce convincing results.
We did many experiments where we stained the cells we were not interested in
and gated them out (we gated out all but B-cells). Although laborious you
could also sort the responding cells (or non responders) and phenotype them
afterwards, thus avoiding all such problems.
Sorry to sound so negative, keep drinking that good beer!
Simon Monard
Aaron Diamond Center
New York
|
|
|
|