Fluorescent In-Situs and FCIS (Vize lab): Difference between revisions
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[[File:33ffar.jpg|200px|thumb| | [[File:33ffar.jpg|200px|thumb|right|FISH, Fluorescein]] | ||
[[File:2876.jpg|200px|thumb| | [[File:2876.jpg|200px|thumb|right|FCIS; fluorescent and colorimetric in situ]] | ||
[[File:38-doubleFISH-big.jpg|200px|thumb| | [[File:38-doubleFISH-big.jpg|200px|thumb|right|Double FISH, Fluorescein and Cy3]] | ||
[[File:frog10x-big.jpg|200px|thumb| | [[File:frog10x-big.jpg|200px|thumb|right|FISH plus transmitted light]] | ||
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 | |||
'''Papers and other sites:''' | |||
Zhou, X. and Vize, P.D. (2004). Proximo-distal specialization of epithelial transport processes within the Xenopus pronephric tubules. [http://www.ncbi.nlm.nih.gov/pubmed/15223337?dopt=Abstract| Developmental Biology 271: 322-338.] | |||
[http://www.faculty.virginia.edu/davidson/fluor_insitu/fluorescent_in_situ.html| Lance Davidson's flourescent in situ methods page] | |||
Revision as of 12:09, 22 December 2009
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
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.