Automated Quantification of Phagocytosis
One of the objectives of NIEHS Center Facility Cores is to develop new methods and technologies for environmental research. The following example is developed by Amy Imrich, Glen DeLoid, Tim Sulahian and Lester Kobzik.
Scanning Cytometry image (A) shows bound (yellow) and internalized (green) fluorescent beads in adherent lung macrophages cultured in 96 well plates. Image analysis software was developed to precisely map beads to cell outlines for counting, shown in (B) as bound (red) vs internalized (green). Hundreds of cells per well in replicate wells are quantitatively analyzed.
High-throughput scanning cytometry assay for lung macrophage phagocytosis. A novel assay has been developed which allows rapid quantitation per cell of fluorescent bead binding and uptake in 96-well plates, facilitating assays of effects of multiple inhibitors, agonists with dose-response and kinetics replicates. This assay has been used to evaluate signaling pathways for the uptake of unopsonized environmental particles, and will allow more mechanistic dissection of how air pollutants cause lung disease. This method was developed using the BioImaging and Flow Cytometry Facilities. A manuscript reporting the findings has been submitted for publication.