Salivary Gland Dissection

Reagend 1:  28 mikrol of 10% NP-40

                   50 mikrol 10X PBS

                   422 mikrol ddH₂O        

Reagend 2:  40 mikrol 10X PBS

                   100 mikrol 16% paraformaldehyde

                   80 mikrol 10% NP-40

                  180 mikrol ddH₂O

Reagend 3: 180 mikrol of 100% acetic acid

                   120 mikrol ddH2O

                   100 mikrol 16% paraformaldehyde

Reagend 4:  450 mikrol of 100% acetic acid

                   550 mikrol ddH₂O

Dissection:

Dissect salivary glands from wondering L3 larvae in 30 mikrol of Reagend 1.

Remove fat body from the glands.

Transfer the glands to mikrol ml Reagend 2, incubate for 40 seconds.

Transfer the glands to mikrol ml Reagend 3, incubate for 40 seconds.

Transfer the glands to 50 mikrol Reagend 4, that is placed in a slide, incubate for 1 min.

Cover with cover slip, gently push with a finger and move in a circle.

Turn the slide upside down, place in a layer of paper tissue, push vigorously.

Place the slide in liquid nitrogen and remove cover slip with scalpel.

Incubate slide in PBST (PBS + 0.1% Tween20)