I performed a phagocytosis assay of PBMC's with K. pneumoniae with moi's of 2:1 and 1:1 for 90, 30 and 15 minutes. The assay was done using 5-DTAF labelled K. pneumoiae and analysed with flow cytometry. And for all three samples this was done for, the percentage of monocytes that phagocytosed decreased as the incubation time increased. Does anyone know of a reason how this could happen?
Joppe Wesseling
Lyudmila Akhmaltdinova
, I added it onto my original post but it as done using 5-DTAF labeled k. pneumoniea, analysed using Flow cytometry. And as experiments with s. marcescens had more than 70% phagocytosis at 90 minutes we used that as a our maximum time point.
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Joppe Wesseling
If someone is looking for the answer here, we discoverd it was because the bacteria were opsonized before being added to the PBMC's. But between the time steps they were kept in the incubator, probably making them more viable for phagocytosis.
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