X-gal staining of fly larvae tissues

1. Dissect fly larvae in PBS.

2. Transfer dissected tissues from several larva to 100 ml of PBS in well slide.

3. Carefully remove PBS with a pipette and add 100 ml PBS/1% Glutaraldehyde. Incubate at room temperature for 15 minutes.

4. Carefully remove PBS/1% Glutaraldehyde and wash twice with 100 ml PBS.

5. Add 50 ml of X-gal staining solution ( 10 mM NaPO4, 150 mM NaCl, 1 mM MgCl2, 6 mM K4[FeII(CN)6], 6 mM K3[FeIII(CN)6], and 0.3% Triton X-100 ) plus 2 mg/ml X-gal. X-gal staining solution should be warmed to 37 C for 5-10 minutes before use.

6. Incubate at room temperature in dark until desired level of staining is observed (several hours to overnight). Stop staining by washing several times with 100 ml PBS.