Reagents and Recipes


Other thoughts: In 'Drosophila Protocols' (Sullivan et al., 2000) it is mentioned that cold methanol during fixation preserves embryo morphology better, and that the formaldehyde concentration should be kept low (1.5-5%) in order to provide the best combination of structural preservation and probe accessibility. After reading many protocols, it seems there is a wide diversity of opinion about what is the best formaldehyde and how much to use. Formaldehyde solution freshly prepared from paraformaldehyde powder is commonly used. I think the bottom line is, the final concentration of formaldehyde in your fix should be about 10% if you want to detect mRNA and 5% if you want to detect protein and preserve ultrastructure as well, and however it gets there it should be good and fresh for killer stains and nice embryo morphology. I'm using Recipe 2 right now, but Recipe 1 works fine.

Embryo pre-treatment and hybridization:

Reagents for making RNA probes:

Reagents for detecting probes:

Mounting media:

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