#  Methods 

 



##  Methods 

 

 

       ![SEM of a Fly Eye](/sites/g/files/omnuum4896/files/styles/hwp_21_9__1920x825/public/electron-microscopy/files/sem_fly_eye.jpg?h=e9427da8&itok=eJ_jJGH7) 

 

 



 

 



 

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  [### Chemical Fixation

 ](/chemical-fixation) To be able to view a biological sample in the electron microscope it must first be stabilized or "fixed", preferrably in a way that the ultrastructure of the cells or tissue remain as close to the living material as possible. The choice of fixative... 

 

 

   ![Oocysts TEM](/sites/g/files/omnuum4896/files/styles/hwp_16_9__480x270/public/2024-11/malhar.jpeg?h=fe10a726&itok=KdA7vG_m) 

 



 

 

   [### Embedding in Resin

 ](/embedding-resin) The routine method for examining biological samples in TEM is to embed the material in plastic and cut ultrathin (60-80nm) sections. The choice of resin depends on the type of sample and the purpose of your study. Listed below are some of the more... 

 

 

   ![Zebrafish Tail Muscle](/sites/g/files/omnuum4896/files/styles/hwp_16_9__480x270/public/2024-11/Screenshot%202024-11-18%20at%209.50.26%20AM.png?itok=OA9DbM-p) 

 



 

 

   [### Frozen Sectioning & Immunogold

 ](/frozen-sectioning) Preparation of ultrathin frozen sections (Tokuyasu) Fixation of cells in culture: Attached cells: first remove cells from the dish with 5mM EDTA/PBS using a transfer pipet with a wide opening. (You can use trypsin if you are not interested in surface... 

 

 

   ![Immunogold Labeled Endoplasmic reticulum ](/sites/g/files/omnuum4896/files/styles/hwp_16_9__480x270/public/2024-11/Screenshot%202024-11-15%20at%2012.04.27%20PM.png?itok=t_6JhFxO) 

 



 

 

   [### Negative Staining

 ](/negative-staining) 5µl of the sample is adsorbed for 1 minute to a carbon coated grid that has been made hydrophilic by a 20 second exposure to a glow discharge (25mA). Excess liquid is removed with a filterpaper (Whatman #1), the grid is then floated briefly on a drop of... 

 

 

   ![Negative Stained Bacteria Phage](/sites/g/files/omnuum4896/files/styles/hwp_16_9__480x270/public/electron-microscopy/files/negative_stained_phage.jpg?h=c8d29249&itok=novndINw) 

 



 

 

   [### SEM Preparation

 ](/sem-preparation) Fixation Fixation 1-2 hrs (or overnight at 4C) 2.5% Glutaraldehyde 2.5% Paraformaldehyde in 0.1 M sodium cacodylate buffer (pH 7.4). Rinse 3 X 10 min in 0.1M sodium cacodylate buffer, pH 7.4 Post-Fix 1-2 hrs in 1% Osmium tetroxide in water Rinse 3 X 5 min... 

 

 

   ![SEM of Squid Tentacle Sucker](/sites/g/files/omnuum4896/files/styles/hwp_16_9__480x270/public/electron-microscopy/files/squi_tentacle_sucker.jpg?itok=fMfwTqO8)