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Standard Operating Procedure
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Title:Freeze Drying Leaf Material
Dept:Agronomy
Lab:Agronomy G313, B411, B406
Supervisor:Randy Shoemaker
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1. Ten minutes before use, the refrigeration unit should be turned on. Do not touch the coils without protective
gloves or with wet hands. These are very cold (-70ºc).
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2. Always use protective clothing when performing this procedure. This means wear a laboratory coat, safety
glasses and gloves when freezing samples.
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3. When placing leaves in the liquid N2, do not stick your fingers into the liquid. Again, wear protective gloves.
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4. Be careful when pouring the liquid N2, it could be easily spilled onto your feet. It is also very cold and could
cause frostbite instantly.
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5. When ready to apply vacuum, make sure all of the spigots are turned to vacuum, turn on the vacuum switch,
and wait until the vacuum light turns green before applying any chambers. When applying a chamber, make
sure the lid and the bucket have no cracks on them (if you see any cracks, please discard the item), and also
check for an abundance of vacuum grease. If you can see or smear the grease, it must be wiped off. A small
amount is helpful, but a large amount could cause the gasket to shift and break the vacuum seal hours after
drawing the vacuum.
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6. Make sure the lid and gasket are centered on the bucket, attach a vacuum hose, and turn the spigot to vent.
This will immediately drop the pressure on the gauge, but the light will turn green again if there are no leaks.
You must be sure the light turns green before leaving the freeze-dryer. The lid gasket may come loose, there
may be a leak, or there might be too much liquid nitrogen in the chamber to allow for good vacuum pressure.
If the vacuum pressure light is not green, the pump may be damaged, and the samples will be degraded.
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7. When samples are dry, usually 48 hours for soybean leaves, slowly turn one of the unused spigots to vent,
thus releasing the pressure. When the pressure has dropped to no whistling noise, then the vacuum pump
should be turned off. Then the refrigeration unit should be turned off. Remove samples, and be nice to the
next person by turning all of the spigots back to vacuum.
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8. Unplug the drain, leaving the brass or black stopper (depending on the freeze dryer being used) in a
noticeable place. Put the drain hose into a bucket for the water to run in to. When the coils are completely
frozen over (no air hole can be seen), the freeze dryer must be stopped and thawed. If the samples are not dry,
water can be used to thaw the coils. Be sure the water is cold, and do not fill evaporation chamber over the
vacuum hole. This will cause water to move into the pump.
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