Preparation of cells for live-cell imaging of a FOXO-based AKT kinase translocation reporter by widefield microscopy

This protocol provides detailed instructions for imaging of cells with stable expression of a kinase translocation reporter, alongside a nuclear marker for segmentation. We have used this approach to measure the nuclear-cytoplasmic shuttling of a FOXO-based AKT reporter in both HeLa cells and human induced pluripotent stem cells (iPSCs). Note that we culture our cells without antibiotics except during live-cell imaging as indicated.

For subsequent data analysis, please refer to the pipeline provided on the Open Science Framework under doi: 10.17605/OSF.IO/4F69N.