Novocrania engrailed fluorescent in situ

Began a fluorescent in situ of Novocrania engrailed to better understand the mesodermic expression. The only difference this time was 10 min of half-concentrated proteinase-k (1.25 µL in 5 mL) instead of 5 min.

24/11/13

Diluted new probe and added to the sample.

26/11/13

Wash day went on smoothly. Added 1:100 anti-dig-pod antibody and incubated in the cold room.

27/11/13

Wash off antibody.

  • Developed with TSA Cy5 for 2h.
  • Stopped reaction with 2x 5 min in detergent solution at 62 °C.
  • 1x 5 min in detergent solution at RT.
  • 3x 5 min PTw at RT.
  • Selected 6 embryos of different stages and went through TDE steps until final 97% + DAPI.
  • Letting overnight in cold room.

28/11/13

Crystals have formed and are sticking around the embryos… Mounted a slide and it works, although there is background.

29/11/13

Confocal at 8h!

Nano en TDE 1.lif - Series004 s1 Nano en TDE 1.lif - Series004 s2 Nano en TDE 1.lif - Series004 s3