Fluorescent In-Situs and FCIS (Vize lab)

These pages contain protocols for performing wholemount fluorescent in situs. Instructions for two-color FISH and for a combination of fluorescent and colorimetric in situs we have called FCIS. Data can be imaged with a fluorescence stereoscope or a confocal.

• FISH protocol
• Fluorescein tyramide synthesis
• Cy3 tyramide synthesis
• FCIS protocol

FISH is not as sensitive as standard BM purple in situs, so if you have a very low abundance message it may not be detectable via fluorescence. You can however do a normal purple development for your low expression gene and a fluorescent counterstain- then overlay them to generate a FCIS image. These are both pretty and very good at highlighting quite subtle overlap

Images:

• 
  FISH, Fluorescein
• 
  FCIS; fluorescent and colorimetric in situ
• 
  Double FISH, Fluorescein and Cy3
• 
  FISH plus transmitted light

Papers and other sites:

Zhou, X. and Vize, P.D. (2004). Proximo-distal specialization of epithelial transport processes within the Xenopus pronephric tubules. Developmental Biology 271: 322-338.

Lance Davidson's flourescent in situ methods page