Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Method of storing cells in a viable state
Reexamination Certificate
2001-01-11
2003-10-14
Naff, David M. (Department: 1651)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Method of storing cells in a viable state
C435S243000, C435S260000, C435S366000, C435S431000
Reexamination Certificate
active
06632666
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The invention relates to gel-based compositions for normothermic, hypothermic and cryopreservative transport or storage of plant tissues or cells and animal organs, tissues or cells, the gel-based compositions comprising a cell maintenance and preservation medium and a gelling agent.
2. Description of Related Art
Today, limited normothermic, hypothermic and cryopreservative maintenance and storage of plant or mammalian cells, tissues and organs (biologics) is accomplished in liquid media. Success is limited, in part, due to damage that occurs during shipment (transport), most often associated with mechanical trauma.
Preservation and transport (herein referred to as preservation) of biologics (cells, tissues, and organs) has traditionally been achieved through “suspension” in a liquid preservation medium. These media include, but are not limited to, simple saline solution, cell culture media, and preservation solutions such as University of Wisconsin (UW) solution (VIASPAN®), EURO-COLLINS®, and HYPOTHERMOSOL® (Bio Life Solutions, Inc., Ewing, N.J., USA). Inherent in this liquid preservation approach is that the liquid environment confers no physical support network for the biologic during preservation and transport. Due to this lack of physical support upon preservation, biologics are exposed to numerous physical stresses during storage and shipment. These stresses include, but are not limited to, sedimentation, mechanical “jarring”, compaction in a liquid column, shaking, vibration, shearing forces, ice damage, and the like. As a result of these mechanical stresses plus additional biochemical stresses inherently associated with biologic preservation in liquid, a significant level of cellular death is initiated during and following the preservation interval. Consequently, failure of the biologic ensues due to this preservation-initiated cell death.
SUMMARY OF THE INVENTION
The invention relates to gel-based medium compositions for normothermic, hypothermic or cryopreservative transport and/or storage of plant tissues or cells and animal organs, tissues or cells, the gel-based compositions comprising a cell maintenance and preservation medium and a gelling agent. In particular, mammalian samples, such as human and animal organs, tissues and cells, may be preserved in the inventive gel-based media compositions.
In a preferred embodiment, the cell maintenance and preservation medium is liquid.
In another embodiment, the gel-based medium compositions comprise:
(a) one or more electrolytes selected from the group consisting of potassium ions at a concentration ranging from about 10-145 mM, sodium ions ranging from about 10-120 mM, magnesium ions ranging from about 0.1-10 mM, and calcium ions ranging from about 0.01-1.0 mM;
(b) a macromolecular oncotic agent having a size sufficiently large to limit escape from the circulation system and effective to maintain oncotic pressure equivalent to that of blood plasma and selected from the group consisting of human serum albumin, polysaccharide and colloidal starch;
(c) a biological pH buffer effective under physiological and hypothermic conditions;
(d) a nutritive effective amount of at least one simple sugar;
(e) an impermeant and hydroxyl radical scavenging effective amount of mannitol;
(f) an impermeant anion impermeable to cell membranes and effective to counteract cell swelling during cold exposure, said impermeant ion being at least one member selected from the group consisting of lactobionate, gluconate, citrate and glycerophosphate-like compounds;
(g) a substrate effective for the regeneration of ATP, said substrate being at least one member selected from the group consisting of adenosine, fructose, ribose and adenine;
(h) at least one agent which regulates cellular levels of free radicals; and
(i) at least one gelling agent.
Yet another embodiment of the invention is a method of storing cell samples in a gel-based medium composition, the method comprising:
warming a gel-based medium composition to melt a gelling agent contained therein;
suspending cell samples in a cell preservation solution without a gelling agent;
mixing the suspended cell samples with the warmed gel-based medium composition;
cooling the cell samples to a chilled or frozen state, thereby solidify the gelling agent; and
transferring the chilled or frozen cell samples to a desired storage temperature.
Optionally, the chilled or frozen cell samples may be transported in the gelled state.
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Baust John G.
Baust John M.
Van Buskirk Robert
BioLife Solutions, Inc.
Miele Anthony L.
Naff David M.
Palmer & Dodge LLP
Ware Deborah K
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