Video Segmentation and Tracking

NF-kB and JNK signaling: Segmentation and Tracking in action!

The following video shows NIH3T3 cells (mouse fibroblasts) expressing a nuclear marker in the green channel (H2B-EGFP), the NF-kB transcription factor in the red Channel (p65-DsRed) and a genetically encoded biosensor for JNK activity in the blue channel (JNK KTR-mCerulean3). Upon addition of Interleukin 1Beta, both NF-kB and JNK are activated by the same upstream mechanisms. This results in the nuclear translocation of NF-kB (red) and a cytoplasmic translocation of the JNK KTR (blue) while the nuclear marker H2B (green) stays always nuclear. The video will play 3 times:

  1. Three channels merged in RGB.
  2. Starts with the three merged channels, then shows just the nuclear channel (H2B-EGFP) in gray scale and how we transform it into segmented objects that we can track over time. Cool!! We typically use these objects to calculate nuclear intensity.
  3. Starts with the gray scale movie for the red channel (p65-DsRed) then we overlay theĀ  cytoplasmic ring objects that we will use to quantify cytoplasmic mean intensity. Finally it goes back to display the three merged channels. Enjoy!