A modified cleansing procedure to obtain large freeze‐fracture replicas

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Abstract
SUMMARY: A method for cleaning freeze‐fracture replicas is elaborated which avoids their rolling up and fragmentating during successive steps. Immediately after freeze‐fracturing, the replicated tissue is slowly thawed from 77 K to room temperature on solid methanol. Safe passage through the subsequent cleaning steps is facilitated by using solutions of gradually increasing surface tension. After at least 24 h of fixation in methanol, the tissue is digested in half‐strength bleach containing 5% ethanol. Following further cleaning in full‐strength bleach, the remaining organic material attached to the replica is dissolved overnight at room temperature in 50% saturated sodium hydroxide.