Methods for production and use of substance-loaded erythrocytes (S-IEs) for observation and treatment of microvascular hemodynamics
First Claim
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1. A method of making a freeze-dried composition of erythrocytes comprising indocyanine green (ICG), the method comprising:
- (a) providing erythrocytes in a blood anti-coagulant solution to form an erythrocyte-containing solution;
(b) reducing the osmolality of the erythrocyte-containing solution from step (a) by adding distilled, de-ionized water directly to the erythrocyte-containing solution to reduce the osmolality of the erythrocyte-containing solution to 100-125 mOsm/kg to form a hypotonic solution to cause pores in the erythrocytes to open;
(c) before performing step (d), combining the hypotonic solution from step (b) with ICG and an at least 50 mM saccharide solution comprising a first saccharide to form an ICG-containing solution;
(d) combining the ICG-containing solution from step (c) with a resealing solution having an osmolality of at least 1000 mOsm/kg to increase the osmolality of the ICG-containing solution, thereby causing the pores in the erythrocytes to close and entrap the ICG and the first saccharide within the erythrocytes;
(e) washing the erythrocytes from step (d) in an isotonic saline washing solution comprising a second saccharide to remove extracellular ICG; and
(f) freeze-drying the washed erythrocytes containing the entrapped ICG from step (e).
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Abstract
Disclosed herein are methods, kits, and compositions for medical imaging relating to fluorescent dyes entrapped in erythrocytes. Also disclosed therein are methods, and compositions further comprising erythrocytes entrapping at least one therapeutically active agent, as well as methods for releasing the entrapped therapeutically active agent(s). Disclosed herein also are methods for preparation of the cells entrapping dye and therapeutically active agent(s) in a freeze-dried form that makes them readily available and easy to use in a clinical environment.
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17 Claims
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1. A method of making a freeze-dried composition of erythrocytes comprising indocyanine green (ICG), the method comprising:
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(a) providing erythrocytes in a blood anti-coagulant solution to form an erythrocyte-containing solution; (b) reducing the osmolality of the erythrocyte-containing solution from step (a) by adding distilled, de-ionized water directly to the erythrocyte-containing solution to reduce the osmolality of the erythrocyte-containing solution to 100-125 mOsm/kg to form a hypotonic solution to cause pores in the erythrocytes to open; (c) before performing step (d), combining the hypotonic solution from step (b) with ICG and an at least 50 mM saccharide solution comprising a first saccharide to form an ICG-containing solution; (d) combining the ICG-containing solution from step (c) with a resealing solution having an osmolality of at least 1000 mOsm/kg to increase the osmolality of the ICG-containing solution, thereby causing the pores in the erythrocytes to close and entrap the ICG and the first saccharide within the erythrocytes; (e) washing the erythrocytes from step (d) in an isotonic saline washing solution comprising a second saccharide to remove extracellular ICG; and (f) freeze-drying the washed erythrocytes containing the entrapped ICG from step (e). - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17)
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Specification