Verwerking van insectenpoten voor fluorescentiemicroscopie: een methode om neuromusculaire structuren voor beeldvorming te behouden

- To begin, submerge anesthetized insects in 70% ethanol solution to reduce the hydrophobicity of the cuticle. Then, wash and permeabilize the tissue with phosphate-buffered saline containing a detergent for at least 30 minutes to allow tissue penetration during fixation.

Next, remove the head and abdomen from the thorax. Use forceps to apply gentle pressure to the coxa-thorax junction, and detach the leg. Carefully place the legs in ice-cold 4% paraformaldehyde solution overnight to allow it to penetrate the tissue. Paraformaldehyde fixes tissues by cross-linking proteins.

Then, remove the fixation solution and wash the tissue several times with phosphate-buffered saline containing detergent. Before mounting, place the legs in pure or slightly diluted mounting buffer for at least one day to provide enough time for the buffer to enter the tissue.

Lastly, mount the legs in a drop of mounting medium. Use a spacer between the microscope slide and the coverslip to prevent the specimen from damage, and secure the mount with nail polish.

In the following example, we will see the dissection, fixation, and mounting of Drosophila melanogaster legs.

- Begin by filling the appropriate number