Measurement of Natural Killer Cell Migration in Response to Chemotactic Stimuli
Transcription
Start by adding chemoattractants, such as a chemokine, in a multiwell plate.
Place a transwell permeable chamber of an appropriate pore size into the well. Add natural killer, or NK, cell suspension to the upper chamber and incubate.
A few chemokine molecules diffuse through the membrane and bind to NK cell receptors. This triggers signaling pathways that promote cytoskeletal changes, resulting in directional cell movement toward the higher chemoattractant concentration in the lower chamber.
After incubation, transfer a fixed volume of migrated cells from the lower chamber to a fluorescence-activated cell sorting or FACS tube. Add a predetermined number of fluorescent counting beads and perform FACS-based cell counting.
Using the following formula, determine the absolute number of migrated NK cells in the sample.
The dot plot gives the number of counting beads and cells separated as distinct regions, based on their fluorescence and scatter properties.
