Protocol for Trachea and Eye Embedding in Mice by TEM

This technique is used to study mice's respiratory and ocular systems. This method involves collecting tissue samples from the trachea and eyes of mice and embedding them in a microscopic medium for analysis. Transmission electron microscopy (TEM) allows for high-resolution imaging of the samples, providing detailed information about the structure and function of these systems.

• Prepare the required instruments and fixative beforehand and place them in a 4°C refrigerator for pre-cooling. Both tracheal and ocular microscopic extractions should be rinsed with saline quickly after dissection or put directly into pre-cooled 2.5% glutaraldehyde fixative (shelf-life of 1 month) and kept in 4°C refrigerator, which is generally required to be completed within two minutes.
• Wash with PB 5 times for 10 min, then 1% osmium tetroxide fixed for 1 h.
• Wash with ddH₂O 5 times for 10 min and stain with uranium for 2 h.
• 30% ethanol, 50% ethanol, 70% ethanol, and 90% ethanol are used in sequence for 15 min, while 100% ethanol is used 2 times for 10 min and epoxypropane 2 times for 10 min.
• Use resin/solvent (1:1, ddH₂O), and resin/solvent (3:1) in that order for more than 2 h. At room temperature, 100% resin for more than 2 h or overnight.
• To accelerate the rate of solidification of the resin, the catalyst DMP-30 is added and then embedded. The embedding is carried out in the embedding plate by adding resin resin-containing catalyst to each small hole in the embedding plate, then picking out a small piece of tissue with a toothpick and placing it at one end of the small hole, picking out another piece and placing it at the other end, making a parallel for each one. And then the embedding plate is put into the oven at 60°C for 48 h or more to make the embedding block solidify and form.

Creative Bioarray Relevant Recommendations

• Creative Bioarray has scientists and imaging laboratories to perform the full comprehensive Transmission Electron Microscopy (TEM) and Scanning Transmission Electron Microscopy (STEM) Services for the biological sciences and clinical research.

• The key to the preparation of biological samples for electron microscopy is that the samples must be made to reflect the state they were in when they lived.
• Don't let the tissue block take off liquid during dehydration and soaking and washing to avoid drying out.

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