Using the Retsch MM301 Ball Mill for Cryogenic
Disruption of Yeast Cells
-1-© Dr. Markus Kalkum 2003
Yeast spaghetti are produced by pressing yeast through small holes into a beaker filled with liquid nitrogen. The
holes (~ 1.0mm Ø) are punched into the bottom of a 50mL Falcon tube that contains 10-20mL of pelleted yeast
obtained after centrifugal harvest of a 2L late log. culture. Pressure is employed manually by inserting the piston
of a 60mL disposable syringe into the tube. Yeast spaghettis (treated with protease inhibitors) can be stored at
-80 ° C.
The stainless steal grinding jars, the grinding balls and the storage tube with the
yeast spaghetti are immersed into liquid nitrogen using a thick-walled
polystyrene box. Pre-cooling is finished when the initially vigorously boiling
nitrogen bath becomes calm.
Grinding jars are removed from the bath and emptied. It is important to remove
all liquid nitrogen form the interior of the jars to avoid possible explosions
During the grinding process. Jars, balls and tubes are best handled with gaspipe pliers or crucible tongs. Always
wear cryo-gloves!
Yeast spaghettis (plus inhibitors) and the grinding ball (s)
are transferred into the jars. The jars should be filled
about half with loose spaghettis (~ 5g). After crushing
and compacting the brittle spaghettis with the ball and
with a precooled spatula, the yeast volume should not
extend above one third of the jar volume – otherwise
grinding becomes inefficient.
Disruption of Yeast Cells
-1-© Dr. Markus Kalkum 2003
Yeast spaghetti are produced by pressing yeast through small holes into a beaker filled with liquid nitrogen. The
holes (~ 1.0mm Ø) are punched into the bottom of a 50mL Falcon tube that contains 10-20mL of pelleted yeast
obtained after centrifugal harvest of a 2L late log. culture. Pressure is employed manually by inserting the piston
of a 60mL disposable syringe into the tube. Yeast spaghettis (treated with protease inhibitors) can be stored at
-80 ° C.
The stainless steal grinding jars, the grinding balls and the storage tube with the
yeast spaghetti are immersed into liquid nitrogen using a thick-walled
polystyrene box. Pre-cooling is finished when the initially vigorously boiling
nitrogen bath becomes calm.
Grinding jars are removed from the bath and emptied. It is important to remove
all liquid nitrogen form the interior of the jars to avoid possible explosions
During the grinding process. Jars, balls and tubes are best handled with gaspipe pliers or crucible tongs. Always
wear cryo-gloves!
Yeast spaghettis (plus inhibitors) and the grinding ball (s)
are transferred into the jars. The jars should be filled
about half with loose spaghettis (~ 5g). After crushing
and compacting the brittle spaghettis with the ball and
with a precooled spatula, the yeast volume should not
extend above one third of the jar volume – otherwise
grinding becomes inefficient.
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