X-Message-Number: 8856
Date: Wed, 26 Nov 1997 22:51:55 -0800 (PST)
From: Doug Skrecky <>
Subject: cryopreservation of neurons

Authors
  Kawamoto JC.  Barrett JN.
Title
  Cryopreservation of primary neurons for tissue culture.
Source
  Brain Research.  384(1):84-93, 1986 Oct 1.

Abstract
  We have developed new cryopreservation methods which allow
  storage of fetal rat central nervous system tissues for more than 1 week at
  3-8 degrees C or for several months at -70 or -90 degrees C prior to tissue
  culture. For refrigeration, small brain regions (less than 2 mm thick) were
  placed intact into 35 mm petri dishes of 'hibernation medium' inside a
  humidified chamber. Optimal preservation was obtained with hibernation media
  of pH 6.8-7.4, containing 30-70 mM K+, 10-30 mM Na+, 5-50 mM PO4(2-), 20 mM
  lactic acid, 5 mM glucose, and less than 0.1 mM Ca2+. The media were made
  approximately isotonic by addition of sorbitol. For
  freezing, brain tissues were dissociated by gentle trituration (without
  enzymes) in the above medium supplemented with 5-10% dimethylsulfoxide. After
  refrigeration or freezing, neurons were very sensitive to damage from
  mechanical stress (e.g. centrifugation, harsh rinsing or trituration). Rapid
  changes in osmotic pressure or excessive polylysine on the tissue culture
  substratum also reduced neuronal survival after
  cryopreservation. Pretreatment of tissue culture substrata
  with media from lung cell cultures, or plating of neurons at higher density
  (2000 cells/mm2) improved neuronal survival after
  cryopreservation.

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