Embryo collection - Acrylic resin

Kidneys were extracted from the embryo, fixed, treated and set in acrylic resin.

Protocol

Fixation

The mother was anaesthetised then perfusion fixed with fresh 4% paraformaldehyde in PBS at 4^oC. The embryos were removed and placed in fresh fix overnight at 4% on a roller. Kidneys were dissected out with adrenal glands still attached. Perfusion fixation is probably not required for embryonic kidneys so they can be dissected fresh from the embryos and immersion fixed.

Dehydration and infiltration

Once fixation was completed, the kidneys were dehydrated in a series of ethanol concentrations up to 70% ethanol (1hr per change) and stored at 4^oC. Prior to embedding the tissues were further dehydrated in a series up to 100% ethanol ( 3x1hr minimum) then placed in LR White acrylic resin overnight at 4^oC.Next day the LRWhite resin was replaced with fresh resin and the kidneys left overnight at 4^oC.

Embedding

A kidney was embedded in the resin plus accelerator in a flat-bottomed foil dish allowed settle. The resin block was kept on ice overnight until it had completely polymerised. The next day the kidney was cut out with a small saw and the block trimmed of excess resin with a single-edged razor blade. The block was then photographed.

Sectioning

The embryos were then subject to histology sectioning