Preparation of Tissue Fixation Embedding and Sectioning

1. Fix tissue in 4% paraformaldehyde in PBS at 4°C overnight (or 4 h at room temperature).

2. Successively replace solution with the following (at least 30 min each with occasional agitation, at room temperature except where stated).

c. 70% Ethanol twice.

f. 100% Ethanol twice.

g. Histolene twice.

h. 1:1 Histolene:melted paraffin wax for 20 min at 60°C.

3. Transfer tissues to a mould at 60°C, orientate and allow wax to set.

4. Cut ribbons of 6 |im sections on a microtome. Float the ribbons of sections on a bath of distilled water at 40°C until free from creases. Collect on subbed slides. Dry slides at 37°C overnight.

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