{"id":294712,"date":"2019-10-04T10:29:10","date_gmt":"2019-10-04T09:29:10","guid":{"rendered":"https:\/\/www.deltamicroscopies.com\/produit\/ems-freeze-substitution-kit\/"},"modified":"2024-02-07T21:04:00","modified_gmt":"2024-02-07T20:04:00","slug":"ems-freeze-substitution-kit","status":"publish","type":"product","link":"https:\/\/www.deltamicroscopies.com\/en\/products\/ems-freeze-substitution-kit\/","title":{"rendered":"EMS Freeze Substitution Kit"},"content":{"rendered":"
\u00a0Kit Features
\n–\u00a0 111 Platform Shaker
\n–\u00a0 002 Ice Chest
\n–\u00a0 003 Heater Block
\n–\u00a0 004 Temperature Probe
\n–\u00a0 005 Cryo Tubes
\n–\u00a0 006 Data Logger<\/div>\n
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Process of Freeze Substitution<\/div>\n
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  1. Cool the metal block by submerging completely in liquid nitrogen in the ice bucket. Leave for 5 minutes or until the “boiling” stops.<\/li>\n
  2. In a separate box, transfer samples into cryovials with frozen fixative, keeping everything at liquid nitrogen temperatures. Seal tightly and be very sure that there is no liquid nitrogen trapped in the vial. Trapped liquid can cause the vials to explode upon warming. It is best to use a room temperature lid when sealing the tube.<\/li>\n
  3. Put the vials with samples into holes in the cooled metal block.<\/li>\n
  4. Go to a PC (Macs won’t work) that has the Lascar datalogger software installed and name and start the program.<\/li>\n
  5. Pour off the liquid nitrogen from the block and box, making sure not to let the cryovials come out of the holes in the block.<\/li>\n
  6. Arrange the block so the cryovials are horizontal and put the tops of the vials against one side of the foam box. Use a piece of foam or wadded up paper behind the block so it keeps the vials from falling out of the block during shaking.<\/li>\n
  7. Turn on the shaker at 100-125 rpm and allow it to gradually warm until the temperature is at least 0\u00b0C before removing the vials for rinsing and resin infiltration. This operation should take place in a fume hood in case there is any leakage of osmium-acetone from poorly sealed vials. Freeze substitution will take about 2 hours with the lid off, and about 3 hours with the lid on (though this may vary from lab to lab).<\/li>\n
  8. Remove the vials from the metal block and place them onto a rocker at room temperature and wait until they come to about 20\u00b0C, then stop the datalogger and save the files.
    \nRinse out the fixative with 3-4 rinses in acetone and proceed to infiltration and embedding. Take care opening the vials as pressure built up inside can cause a spray of the freeze substitution media.<\/li>\n<\/ol>\n

     <\/p>\n

    **NOTE: No dry ice is required for this procedure<\/p>\n\n\n\n\n
    \"\"<\/a><\/td>\n\"\"<\/a><\/td>\n<\/tr>\n
    Tobacco leaf prepared by the quick FS method (Webb & McDonald, 2011). Abbreviations: N = nucleus; Ch = chloroplast; CW = cell wall; Cy = cytoplasm; ER = endoplasmic reticulum; V = vacuole; and, G = Golgi apparatus. Bar = 0.5 \u00b5m.<\/span><\/td>\nTobacco leaf prepared as above showing details of the cell wall (CW) and chloroplast (Ch) and cell membranes. Bar = 100 nm.<\/span><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

     <\/p>\n

    <\/div>\n
    \nWhy does it work?<\/strong><\/div>\n
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