Abstract: Cryopreserved boar spermatozoa is much less fertile than that of other species, which could be due to damage to sperm membrane lipids during the cryopreservation process. Incorporation of selected lipids improves the survival of boar spermatozoa following cryopreservation. Liposomes were made from lipids extracted from head plasma membrane (HPM) of boar spematozoa or from selected lipids (SL) which contained specific phospholipids. At a fixed lipid concentration, fusion efficiency with spermatozoa as measured by flow cytometry and R18 dequenching was affected by lipid type, sperm concentration and incubation time. SL and HPM improved sperm viability (SYBR-14 and propidium iodide) and motility during cooling to 5C., with SL±egg yolk better than or equal to HPM (P<0.05). Post-thaw, egg yolk showed a strong cryoprotective effect.

Abstract: A cryoprotectant solution used for preserving biological material comprising cells is disclosed. The solution comprises dimethyl sulfoxide, an amide such as formamide, urea, acetamide, hydroxyurea, N-methyl formamide, and ethylene glycol or ethylene glycol in combination with propylene glycol wherein the propylene glycol replaces less than 8% w/v of the ethylene glycol.

Abstract: The method of preservation by vitrification, described in the present application, provides for storage of samples at higher temperatures than in conventional methods and can be applied to cells, multicellular tissues, organs and organismes. The method of the present invention includes preparing a solution of vitrification non-permeating co-solutes (amino acids, betaines, carbohydrates, or other non-permeating co-solutes that effectively decrease the chemical potential of permeating cryoprotectants in aqueous solutions), a permeating cryoprotectant and a non-permeating cryoprotectant (polyvinylpyrrolidone, polyethylene glycol, dextran, hydroxy ethyl starch, Ficol, etc.), contacting a sample with the vitrification solution and storing the sample at a storage temperature. The method also includes the step of rehydrating the preserved sample in a rehydration solution prepared in the manner of the vitrification storage solution.

Abstract: This invention relates generally to cryopreservation and a method for preventing injury caused by cooling and warming of tissue and for reducing the toxicity of vitrification solutions. The present method achieves reduction or elimination of injury by increasing the tonicity of the medium to greater than isotonic prior to cooling. The method was developed by attempting to simulate without freezing, the events that take place during freezing living cells and/or tissue. A further benefit of the method is that, since the cryopreservation medium is hypertonic, it can be diluted to a more extreme degree in one step once the system is rewarmed, without engendering the degree of cell swelling that would attend the same dilution factor when diluting an isotonic cryopreservation medium.

Abstract: The instant invention is a novel buffer which enables simultaneous cryoprotection and transfection of mammalian cells. It enables the user to make cell stocks which can be kept long term. Use of these cell stocks circumvents the need to culture cells each time a transfection is undertaken. It alleviates the need for continuous cell culture and repeated transfections due to transfection variability.

Abstract: The present invention provides solutions and methods for preserving living biological materials that enable organs, tissues and cells to be stored for extended periods of time with minimal loss of biological activity. The inventive solutions are substantially isotonic with the biological material to be preserved and are substantially free of dihydrogen phosphate, bicarbonate, nitrate, bisulfate and iodide. In one embodiment, preferred for the preservation of platelets, the solutions comprise betaine, sodium chloride and sodium citrate. For the preservation of many living biological materials, the inventive solutions preferably contain a calcium salt selected from the group consisting of calcium sulfate and calcium chloride.

Abstract: A two stage method of thawing cells from a cryopreserved state includes first warming the cells from a cryopreservation temperature to a transition temperature of at least −30° C. in a first, slow-warming stage by exposing the cells to an atmosphere having a temperature of less than 30° C., and once the cells have obtained the transition temperature, subsequently further warming the cells from the transition temperature by exposing the cells to a temperature of at least 32° C. in a second, rapid-warming stage. After the cells obtain the transition temperature in the first stage, the cells may be equilibrated at the transition temperature for a period of time prior to conducting the second stage warming. The method is particularly useful in warming cryopreserved cells attached to a fixed substrate. A thermal conduction device in association with the cryopreserved cells may also be used to further assist in the warming procedure.

Abstract: A new method of cryopreservation based on the very fast cooling rates achieved by the direct contact of small droplets of vitrification solution containing biological sample with a very cold solid surface.

Abstract: Disclosed is a biopharmaceutical product cryopreservation system, for cryopreserving a biopharmaceutical product that includes a cryopreservation compartment; a cryopreservation fluid located within the cryopreservation compartment; and a biopharmaceutical product cryopreservation vial located within the cryopreservation compartment and surrounded by the cryopreservation fluid, and the biopharmaceutical product cryopreservation vial including a body that includes an oblong cross-section defining proximal and distal ends of the body, and at least one nucleating structure, coupled to a distal end of the body, and the body including a cryogenically stable material that is compatible with biopharmaceutical products. Also disclosed are cryopreservation vials and methods of cryopreserving biopharmaceutical products.

Abstract: A method is provided for freeze-drying multiple samples of viable microorganisms which method comprises: (i) providing a container comprising multiple wells; (ii) dispensing multiple liquid samples, each sample comprising a viable microorganism, into separate wells of the container; (iii) placing the container in a freeze-drying apparatus; and (iv) freeze-drying the samples present in the wells under conditions that substantially maintain the viability of the microorganisms. Also provided is a container comprising multiple wells, each well comprising a viable freeze-dried sample of a microorganism.

Abstract: A method for cooling, rewarming, and removing cryopreservative of living cells by decreasing injury and cryotoxicity is presented. The method was developed by attempting to simulate without freezing, the events that take place during freezing living cells and/or tissue.

Abstract: The invention relates to the field of organ and tissue perfusion. More particularly, the present invention relates to a method for preparing organs, such as the kidney and liver, for cryopreservation through the introduction of vitrifiable concentrations of cryoprotectant into them. To prepare the organ for cryopreservation, the donor human or animal, is treated in the usual manner and may also be treated with iloprost, or other vasodilators, and/or transforming growth factor &bgr;1. Alternatively, or additionally, the organ which is to be cryopreserved can be administered iloprost, or other vasodilators, and/or transforming growth factor &bgr;1 directly into its artery. The invention also relates to preparing organs for transplantation by a method for the removal of the cryoprotectant therefrom using low (such as raffinose, sucrose, mannitol, etc.

Abstract: An apparatus for freezing live cells, in particular sperm, which cells are included in at least one sample in at least one container, wherein means are provided for cooling the or each container, wherein the cooling means (5, 6, 7; 105, 106, 107) comprise at least one contact face (3, 103), capable of being cooled, for cooling, during use, the or each container (16, 116) and the or each sample included therein through abutting contact, wherein control means (10, 110) are provided for controlling, during cooling, the cooling rate (TG) and the ambient temperature of the or each container (16, 116).

Abstract: A method for preserving biological material includes the steps of placing the biological material in thermal contact with a cryogenically coolable environment, applying radiant energy to the biological material to maintain the temperature of the biological material at physiological temperatures, cooling the surrounding environment to a temperature below the glass phase transition temperature of the biological material, and rapidly stopping the application of radiant energy to the biological material. The method produces cooling rates so rapid that the biological material is vitrified without an opportunity for ice crystals to form.

Abstract: The invention relates to cells or tissues having an increased amount of regulatory proteins, including cytokines, growth factors, angiogenic factors and/or stress proteins, and methods of producing and using those cells or tissues. The invention is based on the discovery that the production of regulatory proteins is induced in cells or tissue constructs following cryopreservation and subsequent thawing of the cells or constructs. The compositions and methods of this invention are useful for the treatment of wound healing and the repair and/or regeneration of other tissue defects including those of skin, cartilage, bone, and vascular tissue as well as for enhancing the culture and/or differentiation of cells and tissues in vitro.

Abstract: Methods of fixing and processing tissue and samples on a membrane by using ultrasound radiation as a part of the method are presented. Ultrasound of a frequency in the range of 0.1-50 MHz is used and the sample or tissue receives 0.1-200 W/cm2 of ultrasound intensity. The use of ultrasound allows much shorter times in the methods. Also presented are apparati comprising transducers of one or of multiple heads for producing the ultrasound radiation and further comprising a central processing unit and optionally comprising one or more sensors. Sensors can include those to measure and monitor ultrasound and temperature. This monitoring system allows one to achieve accurate and optimum tissue fixation and processing without overfixation and tissue damage. The system also allows the performance of antigen-antibody reactions or nucleic acid hybridizations to be completed in a very short time while being highly specific and with a very low or no background.

Abstract: A process for preparing pliable soft tissue specimen which are resistant to cracking and devoid of viable cells includes the steps of treating native soft tissue obtained from a donor by a gradually increasing gradient of aliphatic alcohol or other suitable water miscible polar organic solvent until the last alcohol (or other solvent) solution has at least 25% by volume of the organic liquid. Thereafter, the tissue specimen is treated with a solution containing glycerol or low molecular weight (<1000 D) polyethylene glycol, and polyethylene glycol of a molecular weight between approximately 6,000 to 15,000 D and heparin. Thereafter, the tissue specimen is briefly immersed in aqueous heparin solution, frozen and lyophilized. The tissue specimen is suitable for implantation as a homograft or xenograft, with or without rehydration.

Abstract: A method of cooling and rewarming large organs and tissues at sub-zero temperatures is provided. The method includes perfusion of tissue with inert fluids that remain liquid at very low temperatures. The inert fluids can include fluorocarbons, silicones, or fluorosilicones. Glycol ethers are used as a novel class of cryoprotective agents for organ and tissue preservation. The glycol ethers have improved viscosity, penetration, and glass forming properties.

Abstract: A method for removing a target DNA fragment having a predetermined base-pair length from a mixture of DNA fragments comprises the following steps. A mixture of DNA fragments which may contain the target DNA fragments is applied to a separation column containing media having a nonpolar, nonporous surface, the mixture of DNA fragments being in a first solvent mixture containing a counterion and a DNA binding concentration of driving solvent in a cosolvent. The target DNA fragments are separated from the media by contacting it with a second solvent solution containing a counterion and a concentration of driving solvent in cosolvent which has been predetermined to remove DNA fragments having the target DNA fragment base pair length from the media. The target DNA fragments can be collected and optionally amplified. When the method is being applied to collect a putative fragment, if present, no DNA fragments having the base pair length of the target DNA could be present in the mixture.

Abstract: Methods for labeling mannose lectins on the surface of mammalian sperm cells are described, and methods of categorizing and quantifying the numbers of cells exhibiting particular labeling patterns are also provided. These methods provide ways of detecting whether or not a mammalian male patient has mannose lectin-correlated infertility, which is determined on the basis of the relative numbers of sperm cells labeled with particular labeling patterns. Methods for detecting capacitation-induced changes in the distribution of mannose lectins on mammalian sperm cells are also provided, and these methods can also be used to detect whether or not a mammalian male patient has mannose lectin-correlated infertility, on the basis of the comparative relative labeling in capacitated and non-capacitated sperm cell samples. In addition, the various methods are shown to be useful for determining whether chemical compounds have effects on mammalian sperm cell surface lectin distribution or acrosomal states.

Abstract: The present invention relates to a medium allowing the preservation and cryopreservation of biological materials such as animal cells and viral particles that are directly injectable or reinjectable into an organism. A medium for preserving and/or freezing biological materials, including a saline solution, modified fluid gelatin and human serum albumin, is disclosed.

Abstract: Improved artificial diets or growth media are described which are suitable for rearing large numbers of viable and biologically fit arthropods, including zoophagous arthropods and phytophagous arthropods, including facultatively zoophagous arthropods. In a first embodiment, the growth medium is composed of a mixture of cooked egg, liquid, and carbohydrate source. In a second embodiment, the growth medium is composed of a plant-based phytophage diet which includes cooked egg yolk or cooked whole egg. In a third embodiment, the growth medium is composed of a mixture of cooked egg, liquid, and carbohydrate source in admixture with a plant-based phytophage diet which includes cooked egg yolk or cooked whole egg. The growth media are devoid of meat products or insect components and are suitable for mass production of arthropods at a reasonable cost for use in biological control programs or other biologically based technologies.

Abstract: This invention relates to methods, apparatus and solutions for cryopreserving microscopic biological materials for biologically extended periods of time. The method comprises treating a suspension of biological material, in an appropriate buffer, with a cryoprotectant or combination of cryoprotectants which raises the glass transition temperature range of the sample. One or more dry protectants may be added to the cryosolution. The cryosolution is then nebulized and rapidly cooled with novel apparatus, dried by molecular distillation, stored and then rehydrated in a buffer prior to its use. The solutions comprise novel mixtures of cryoprotectants.

Abstract: A method for vitrification of a blood vessel includes immersing the blood vessel in increasing concentrations of cryoprotectant solution at a temperature greater than −15° C. to a cryoprotectant concentration sufficient for vitrification; cooling the blood vessel at an average rate of from 30-60° C. per minute to a temperature between −80° C. and the glass transition temperature; and further cooling the blood vessel at an average rate less than 10° C. per minute to a temperature below the glass transition temperature to vitrify the blood vessel. After the vitrified blood vessel has been stored, the blood vessel may be removed from vitrification by warming the blood vessel at an average rate of from 20-40° C. per minute to a temperature between −80° C. and the glass transition temperature; further warming the blood vessel at a rate of from 200-300° C. per minute to a temperature above −15° C.; and reducing the concentration of the cryoprotectant.

Abstract: The invention relates to the field of organ and tissue perfusion. More particularly, the present invention relates to a method for preparing organs, such as the kidney and liver, for cryopreservation through the introduction of vitrifiable concentrations of cryoprotectant into them. To prepare the organ for cryopreservation, the donor human or animal, is treated in the usual manner and may also be treated with iloprost, or other vasodilators, and/or transforming growth factor &bgr;1. Alternatively, or additionally, the organ which is to be cryopreserved can be administered iloprost, or other vasodilators, and/or transforming growth factor &bgr;1 directly into its artery. The invention also relates to preparing organs for transplantation by a method for the removal of the cryoprotectant therefrom using low (such as raffinose, sucrose, mannitol, etc.

Abstract: The invention provides a method of isolating cells from a faecal stool, the method comprising the steps of a) cooling the stool to a temperature below its gel freezing point, and b) removing cells from the stool whilst maintaining the stool at a temperature below its gel freezing point such that the stool remains substantially intact. The invention further provides methods of purifying cells comprising use of immunomagnetic beads and/or boric acid. Cells isolated according to the invention may be used in diagnostic tests and assay procedures for monitoring a biological or biochemical property of tissue.

Abstract: The present invention provides methods and compositions for cryopreservation of cells or tissues encapsulated in a bioartificial organs, wherein the integrity and the viability of the encased cells are maintained, along with the integrity of the artificial capsule used to encase the cells. The method provides novel conditions for cryopreserving a bioartificial organ in a freezing container using a minimum volume of added cryopreservative solution necessary to maintain both the structural integrity of the jacket encasing the bioartificial organ, and the viability of the cells encased therein.