Cryopreservation of rat and human liver slices by rapid freezing.
The cryopreservation of human liver slices is a promising way to enhance the ability to test the metabolism of drug candidates. This study demonstrates the use of a novel technique for the cryopreservation of both rat and human liver slices. In this technique the slices are treated with Me2SO and sandwiched between aluminum plates separated by a thin gasket. The device is then submerged in liquid nitrogen to freeze the slices, which can then be stored until use. To thaw the slices, the apparatus is submerged in a water bath at 37 degrees C. Slices frozen and thawed in this manner were compared to those frozen in conventional cryovials. The viability of the slices was determined by incubating them in 12-well plates and measuring urea synthesis, ethoxycoumarin metabolism, and cytosolic enzyme leakage (LDH and ALT). The viability of rat slices frozen between plates approached that of fresh slices and was consistently higher than slices frozen in cryovials. Slices from two human samples gave similar results. The technique was found to work over a wide range of Me2SO concentrations (4.5 to 22% was tested) with an optimal concentration between 10 and 15%.
Day
S H
SH
Merck Frosst Centre for Therapeutic Research, Pointe-Claire-Dorval, Quebec, H9R 4P8, Canada.
Nicoll-Griffith
D A
DA
Silva
J M
JM
D000368:Aged; D000410:Alanine Transaminase / Q000378:metabolism; D000818:Animals; D003374:Coumarins / Q000378:metabolism; D015925:Cryopreservation / Q000295:instrumentation / Q000379:methods*; D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide; D005069:Evaluation Studies as Topic; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009926:Organ Preservation / Q000295:instrumentation / Q000379:methods*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013045:Species Specificity; D014508:Urea / Q000378:metabolism
D003374:Coumarins; D003451:Cryoprotective Agents; C017299:7-ethoxycoumarin; D014508:Urea; D007770:L-Lactate Dehydrogenase; D000410:Alanine Transaminase; D004121:Dimethyl Sulfoxide