Hi all-
What would be a good positive controls for phagocytosis/pinocytosis for color-based flow cytometry assay in antigen presenting cells?
For testing pinocytosis fluorescence labeled dextran is the best option.
For testing phagocytosis fluorescent labeled particles or bacteria.
Hi,
I am not sure FITC dextrane is a good option for you as it is believed that FD is taken up by receptor mediated endocytosis.
Many people use Lucifer Yellow for measuring pinocytosis. My recommendations would be:
1. For phagocytosis see the previos reply. You might also want to try adding dead cells to MFs/phagocytic cells, in some context this type of readout could be more "physiological". Working with latex beads is surely easy and fast, but does not report a lot of aspects of phagocytosis (e.g. the metabolic burden of eaten material).
2. For pinocytosis, use LY
3. For receptor mediated endocytosis, use FITC Dextrane. I previously used the Mw 70K variety.
I have to add that in these experiments (pino/receptor med endo/phagocytosis) your readout depends a lot on cell types/substrates/condition/incubation length (!) etc and therefore it is very important to try different conditions, then choose one and standardize that rigorously.
Many people think that all these experiments give you limited insight into "true" phagocytic capacities. I do not know to what extent to agree with them, but all these phagocytosis experiments have a tendency to show large lab-to-lab variability.
T
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