Analyzing Cell Migration via Folate Chemotaxis

Cellular morphology, pseudopod dynamics and cell migration were measured by an under-agarose folate chemotaxis assay as described previously (Singh et al., 2020 (link); Singh and Insall, 2022 (link)). In brief, 0.4% SeamKem GTG agarose in Lo-Flo medium (Formedium) was boiled. After cooling, folic acid to a final concentration of 10 μM was added. 5 ml of agarose-folate mix was poured into the 1% BSA-coated 50 mm glass bottom dishes (MatTek). A 5 mm wide well was cut with a sterile scalpel and filled with 200 μL of 2 × 10⁶ cells/ml. Cell migration was imaged after 4–6 h with 10x and 60x differential interference contrast (DIC) microscopy. To examine the localization of labelled proteins in the pseudopods, cells were also imaged by AiryScan confocal microscopy.