WO2025257595A1 - Solution de conservation - Google Patents

Solution de conservation

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Publication number
WO2025257595A1
WO2025257595A1 PCT/IB2024/055741 IB2024055741W WO2025257595A1 WO 2025257595 A1 WO2025257595 A1 WO 2025257595A1 IB 2024055741 W IB2024055741 W IB 2024055741W WO 2025257595 A1 WO2025257595 A1 WO 2025257595A1
Authority
WO
WIPO (PCT)
Prior art keywords
preserving
biological tissue
preparation
tissue
solution
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/IB2024/055741
Other languages
English (en)
Inventor
Filippo Naso
Alessandro Gandaglia
Ugo STEFANELLI
Claudio GATTO
Jana D'AMATO TOTHOVA
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Biocompatibility Innovation Srl
Original Assignee
Biocompatibility Innovation Srl
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Biocompatibility Innovation Srl filed Critical Biocompatibility Innovation Srl
Priority to PCT/IB2024/055741 priority Critical patent/WO2025257595A1/fr
Publication of WO2025257595A1 publication Critical patent/WO2025257595A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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Classifications

    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/10—Preservation of living parts
    • A01N1/12—Chemical aspects of preservation
    • A01N1/122—Preservation or perfusion media
    • A01N1/126—Physiologically active agents, e.g. antioxidants or nutrients

Definitions

  • BPs bioprostheses
  • These biological substitutes often derived from animal or human tissues, represent a paradigm shift in medical interventions, particularly in the fields of cardiac, orthopedic, and vascular surgeries aimed at the replacement or augmentation of damaged or diseased tissues within the human body.
  • BPs alternatively known as biological or tissue-based prosthetic devices, embody a diverse array of substitutes designed.
  • they offer a remarkable advantage in terms of biocompatibility and functional mimicry of natural tissues.
  • the rationale behind their development is to create interventions that seamlessly integrate with the body's biological processes, promoting improved patient outcomes while minimizing the risk of complications.
  • BPs are crafted from a variety of biological materials, with xenografts and allografts being the predominant sources.
  • Xenografts derived from animal tissues, often include porcine or bovine tissue-derived material, chosen for their anatomical and physiological similarities to humans. Allografts, sourced from human donors, offer the advantage of reduced immunogenicity but pose challenges related to availability and potential disease transmission.
  • tendons and ligaments of bovine origin can be used as a biological substitute for the compromised human structure since they show good biocompatibility, and strength as well as lack adhesion to surrounding structures.
  • Corneas are a common allograft BPs. This tissue can be stored in appropriate banks and used after carefully verifying compatibility between recipient and donor [1 ].
  • Vascular BPs often derived from xenogeneic sources, address the challenges of vascular grafting by providing conduits that integrate seamlessly with the host vasculature, minimizing the risk of thrombosis and graft failure [2].
  • BPs While the biological nature of BPs offers advantages in terms of compatibility, it introduces challenges related to their preservation.
  • the success of these implants hinges on the ability to maintain the structural and functional integrity of the biological tissues over time.
  • the inherent susceptibility to enzymatic degradation, immunogenic responses, and other biological processes necessitate optimal preservation methods to ensure the sustained efficacy of bioprosthetic devices in the long term.
  • cryopreservation and chemical fixation have been the cornerstones of BPs preservation.
  • the former relies on low temperatures to halt biological processes and extend the viability of tissues. This method is particularly relevant for allografts, allowing for long-term storage and convenient distribution.
  • cryopreservation introduces challenges related to ice crystal formation, which can damage cellular structures and compromise the functional properties of the bioprosthetic material upon thawing [3].
  • GA is responsible for the incomplete masking of the xenogeneic antigens present on the surface of the animal tissues used for the manufacture of BPs.
  • GA is still used in the manufacture of BPs and the preparation of the storage solution adopted for their packaging. These medical devices are supplied in a 0.2% GA buffered saline solution, which helps maintain sterility and tissue stability.
  • Implantable medical devices that are beginning to break away from the use of GA as a preservation medium have appeared on the market in the recent period, but they are still very rare. These include, for example, the Inspiris Resilia BHV (Edwards Lifesciences, USA) [7]. Another example is represented by Corcym’s Perceval BHV which is preserved in a buffered aldehyde-free sterile solution that doesn’t need to be rinsed before used and it’s claimed to be free from free aldehyde [8]. Also, other types of implantable devices than BHVs, such as patches for reparative surgery made of equine pericardium or porcine submucosa, are distancing themselves from the use of GA. For example, Matrix Patch (Tissue Berlin) is treated without GA, stored, and delivered in an antibiotic preservative solution. Cardiocel patch (LeMaitre) is also sterilized and stored with a technology that does not involve the use of aldehyde.
  • Matrix Patch T
  • the ideal storage solution should be nontoxic for the patient and able to preserve the BPs for a long time.
  • the solution containing the BPs needs to be stable over a wide temperature range for easy transport and storage. It should also protect from microbial contamination as any infection can lead to thrombosis and degeneration of the device. Additionally, it should support the maintenance of a non-oxidative environment. The oxidation of collagen, the main component of BPs, leads to its fragmentation, making the tissue more susceptible to degradation by proteolytic enzymes and impairing the mechanical performance of the device [12].
  • the optimal storage solution for an implantable medical device should be aldehyde-free, bacteriostatic, non-oxidative, stable over time, and harmless to patients.
  • This preservative solution is based on the synergic action of polyol molecules, sulfur acid salts, and a peptide with antioxidative properties.
  • polyols are monomers or polymer organic compounds characterized by the presence of multiple hydroxyl groups; based on the number of hydroxyl groups these compounds can be divided into diols, triols, and tetrols when bearing two, three, and four hydroxyl groups respectively.
  • Being hydrogenated carbohydrates polyols find wide use in both food and pharmaceutical preparations.
  • An example of monomeric polyol is glycerol which is a trivalent alcohol normally present in human tissue, where it forms the skeleton of the triglycerides, among other functions. It is a colorless, viscous liquid, stable under most conditions. Glycerol is nontoxic, easily digested, and is environmentally safe.
  • Polymeric polyols are subdivided into polyethers, polyesters, and polycarbonates.
  • Polyether polyols can be further subdivided into polyethylene oxide or polyethylene glycol (PEG).
  • PEG is a petroleum-derived polyether compound that finds diverse applications, ranging from industrial manufacturing to medicinal uses. It is used as an excipient in parenteral and ophthalmic formulations since it makes a viscous solution and prevents active ingredients from flocculating and as a lubricant in tablets helping the flow of powders.
  • sulfites are salts derived from sulfurous acid and encompass substances such as sodium sulfite, potassium sulfite, and calcium sulfite that are used as food additives and preservatives in wine, juices, and meat.
  • hydrogen sulfites distinguished by the HSOs- anion; an example is sodium hydrogen sulfite, commonly referred to as sodium bisulfite with the same uses described above for the other salts.
  • the group of sulfurous acid salts comprises metabisulfite too.
  • Chemical compounds present the S2Os 2- ion which salts, once dissolved in water, release the HSOs- anion. These include sodium metabisulfite which besides its use in food and beverages as a preservative, is used as an excipient in medications containing adrenaline since being an antioxidant it prevents its oxidation.
  • peptides are a heterogeneous class of chemical compounds made by a chain of amino acids that bind with a carbamide bond. They are subdivided according to the length of the chain into dipeptides, consisting of two amino acids, tripeptides, when the chain contains three amino acids, tetra peptides when it contains four amino acids, and polypeptides when the chains consist of five or more amino acids. Certain peptides possess the ability to influence physiological functions in both human and animal organisms, earning them the designation of bioactive peptides. These peptides can serve various functions, including but not limited to antihypertensive, immunomodulatory, antimicrobial, mineral carrier, and antiviral activities.
  • Glutathione is a compound consisting of glutamic acid attached via its side chain to the N-terminus of cysteynil-g lycine. It has many roles in cells: it is conjugated to drugs to make them more soluble for excretion (glutathione conjugation is an important detoxification mechanism), is a cofactor for some enzymes, is involved in protein disulfide bond rearrangement, and reduces peroxides. Chemically synthesized tetrapeptides are widely used in cosmetics as antioxidant-acting components in facial creams.
  • This invention aims to create a Preserving Solution for bioprosthetic devices (consisting entirely or partly of biological tissue) that does not involve the use of aldehydes (and their derivatives) and to preserve the features of the BPs unchanged over time.
  • the solution must be able to maintain an antioxidant environment, avoiding any degeneration of the preserved BP (such as variation in color or size).
  • Another essential feature is to avoid the growth of microorganisms ensuring the sterility maintenance of the device.
  • the present invention discloses a solution for storing and preserving biological tissues.
  • the present invention discloses a kit for the preparation of a storing solution.
  • the present invention discloses a method for storing and preserving a biological tissue or a portion thereof.
  • the first aspect of the invention provides an organ and biologic tissue-preserving preparation (hereinafter Preserving Preparation) for static storage preservation that demonstrates superior quality preservation when compared to existing preserving media based or not on aldehyde.
  • Such organ and biologic tissue Preserving Preparation includes at least, a polyol molecule, sulfur acid salts, and a peptide with antioxidative properties.
  • the polyol molecules are selected from the group of low molecular weight polyols (glycerol, trimethylolpropane, pentaerythritol, 1 ,4-butanediol, and sugar alcohols such as maltitol, sorbitol, xylitol, erythritol, and isomalt), and polymeric polyols (polyurethanes, polyvinyl alcohol), but preferentially pure glycerol is used.
  • glycerol trimethylolpropane
  • pentaerythritol pentaerythritol
  • 1 ,4-butanediol and sugar alcohols
  • sugar alcohols such as maltitol, sorbitol, xylitol, erythritol, and isomalt
  • polymeric polyols polyurethanes, polyvinyl alcohol
  • pure glycerol is used at a concentration between 1 % and 100% v/v, and preferably at the concentration of 80% v/v.
  • the sulfur acid salts are sodium metabisulfite or sodium pyrosulfite.
  • sodium metabisulfite is used at a concentration between 0.1% and 50% w/v, and preferably at the concentration of 4% w/v.
  • the antioxidative peptide is selected from the group of tripeptides with a gamma peptide linkage between the carboxyl group of the glutamate side chain and cysteine, preferentially glutathione is adopted.
  • glutathione is glutathione-reduced.
  • glutathione-reduced is used at a concentration between 0.05% and 50% w/v, and preferably at the concentration of 1% w/v.
  • the Preserving Preparation can include an apoptosis inhibitor.
  • the apoptosis inhibitor is a caspase inhibitor, a BID inhibitor, a BAD inhibitor, a BAX inhibitor, a BAK inhibitor, a cytochrome C inhibitor, a cathepsin inhibitor, a granzyme B inhibitor, a pyroptosis inhibitor, and/or a necroptosis inhibitor.
  • the caspase inhibitor is a pan-caspase inhibitor.
  • the caspase inhibitor is emricasan (IDN-6556), VX-765 (belnacasan), Q-VD- OPh, VX-166, VX- 740, GS-9540, Ac-DEVD-CHO, Ac-FLTD-CMK, Z-DEVD-FMK, INF 4E, Z-VAD-FMK, or a derivative thereof.
  • the caspase inhibitor is a caspase 3 inhibitor, a caspase 7 inhibitor, a caspase 4 inhibitor, a caspase 8 inhibitor, a caspase 1 inhibitor, a caspase 2 inhibitor, a caspase 6 inhibitor, a caspase 9 inhibitor, or a caspase 10 inhibitor.
  • the Preserving Preparation can further include one or more kinase inhibitors, Rho-kinase inhibitors, polyamines, integrated stress response inhibitors, reactive oxygen species scavengers, and/or antioxidants.
  • one or more agents belonging to the group of protectants, cryoprotectants, ice nucleators, ice modulators, oxygen carrier agents, oncotic agents, crystalloid components, colloidal components, growth factors, vasodilators, kinase inhibitors, Rho-kinase inhibitors, polyamines, integrated stress response inhibitors, reactive oxygen species scavengers, antioxidants, and combinations thereof, can be added to the solution.
  • the Preserving Preparation can include a mixture of antioxidants that reduce reactive oxygen species (ROS), reduce oxidation of lipids, and inhibit or reduce peroxidation.
  • antioxidants that reduce reactive oxygen species (ROS), reduce oxidation of lipids, and inhibit or reduce peroxidation.
  • ROS reactive oxygen species
  • examples include zinc, vitamin C (ascorbic acid), vitamin E (alphatocopherol), selenium, Trolox, ebselen, glutathione, carotenes, ubiquinol (coenzyme Q), propyl gallate (PG), butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), hydrogen sulfide, erythorbate, sodium tripolyphosphate, ethylenediaminetetraacetic acid, ethoxyquin, caseinates, pyruvate and similar compounds that inhibit oxidation.
  • the solution may combine multiple antioxidants for their different properties to prevent oxidation of lipids and accumulation
  • the Preserving Preparation may contain molecules or compounds that inhibit enzymes that metabolize arachidonic acid and other fatty acids that contribute to inflammatory pathways and cell death. This can be selected from the group that includes inhibitors to cyclooxygenase (COX) isoenzymes formally known as prostaglandin-endoperoxide synthase. Examples of inhibitors are nonsteroidal antiinflammatory drugs which include celecoxib and rofecoxib.
  • the Preserving Preparation may include additives that stabilize and maintain cell membrane or protect the cellular membrane from stresses.
  • additives that protect cell membranes include tryptophan, or polyethylene glycol (PEG-35).
  • Other compounds that stabilize the cell membrane include trehalose or other sugars, glycerol, alginate, hyaluronic acid, and dimethyl sulfoxide.
  • the Preserving Preparation can contain oxygen or nitrogen-free radical scavengers including but not limited to N-acetylcysteine (NAC), glutathione, allopurinol, and/or material with redox properties such as cerium oxide nanoparticle, rod, or other forms.
  • the storage solution may include one or a mixture of these compounds.
  • the Preserving Preparation may include a combination of any of the above inhibitors with various functions to inhibit pathways associated with lipotoxicity, ROS, and oxidized lipids.
  • the Preserving Preparation may include antibiotics including gentamycin or vancomycin.
  • the biological tissue sample and recovered biological tissue sample can be fresh, cellularized and viable, cryopreserved, fixed with aldehydes or derivatives, decellularized, crosslinked with polyphenols, resins, and epoxides, or treated with anti-calcific, anti-thrombotic, anti-protein adsorption, anti-bacterial methods or any combination thereof.
  • the biological tissue may consist of a bioprosthetic device, a biopsy specimen, or a portion of a tissue district intended for transplantation (auto- or allo-graft) or analysis.
  • the biological tissue may be of human or animal origin.
  • bioprosthetic tissue includes but is not limited to cardiac valve, tendon, ligament, pericardium, muscular fasciae, dura mater, tympanic membrane, intestinal submucosa, cartilage, adipose and bone tissue, pelvic, abdominal, breast, nerve graft tissue urinary bladder membranes, epineurium, placenta, amniotic membrane, chorionic membrane, umbilical cord tissue, umbilical veins, Wharton’s jelly, and dermal tissue.
  • the disclosed preparation may be in liquid form or in solid form to be reconstituted.
  • the preparation can be supplied in the form of a kit and different bottles containing the correct dosage of powders and liquids for proper reconstitution together with technical instructions.
  • the biological organ and tissue Preserving Preparation can be reconstituted using sterile water for injection, sterile distilled deionized water, sterile water for irrigation, isotonic saline solution, buffered solutions, and physiological solutions.
  • Preferentially sterile water for injection (SWI) is adopted.
  • SWI is used at a concentration between 1 % and 99% v/v, and preferably at the concentration of 20% v/v.
  • the disclosed preserving preparation is in the form of a solution, which is therefore referred to a Preserving Solution.
  • a method for preserving or storing a biological tissue comprising the step of contacting the preparation of the invention with the biological tissue for a sufficient period of time.
  • the biological tissue is the one above disclosed.
  • the preparation is the one above disclosed.
  • the step of contacting may be any one of washing, immersing, submersing, perfusing the biological tissue with the disclosed preparation.
  • the preparation is a solution.
  • the preparation includes a normothermic solution, a sub-normothermic solution, a perfusion solution, or a subzero solution.
  • the time of contacting may be from a few seconds up to a few months.
  • the time of contacting may be of less than 1 minutes to 3 months or even up to 12 months or more.
  • the time of contacting may be up to 5 years or more.
  • said contacting is performed before, during, and/or after cooling the biological tissue sample to a sub-normothermic condition (e.g., from about 12°C to 35°C or any other temperature range described herein), a hypothermic condition (e.g., from about 0°C to 12°C or any other temperature range described herein), or a subzero condition (e.g., below -4°C or any other temperature range described herein).
  • a sub-normothermic condition e.g., from about 12°C to 35°C or any other temperature range described herein
  • a hypothermic condition e.g., from about 0°C to 12°C or any other temperature range described herein
  • a subzero condition e.g., below -4°C or any other temperature range described herein.
  • hypothermic condition or the subzero condition includes hypothermic cooling, supercooling, subzero non-freezing, partial freezing, cryopreservation, or vitrification.
  • Bovine pericardium was obtained from a certified abattoir (Inalca Spa, Castelvetro di Modena, Italy) and transported to the laboratory under controlled conditions.
  • the pericardial samples were harvested from the anterior region of the heart after washing thoroughly in sterile phosphate-buffered saline (PBS) at 4°C.
  • PBS sterile phosphate-buffered saline
  • the most homogeneous portions of pericardial tissue were selected through the use of an optical viewer and the thickness measurement (MTG, Digital Material Thickness Gauge, ELECTROMATIC Equipment Co., New York).
  • the selected pericardium area was mechanically cleaned from adipose tissue debris and dissected.
  • the samples were placed into 12cmxl 0cm bespoke frames ( Figure 1 ).
  • G samples pericardial tissue patches were incubated in a buffered GA solution for three steps of 24 hours each in a dark room.
  • a GA solution is a 0.6% ⁇ 0.5% v/v for the first and second steps followed by a 0.2% ⁇ 0.15% v/v for the third ones.
  • GA- treated pericardial patches were subjected to two washing steps in phosphate buffer for 15 minutes each.
  • GP samples pericardial tissue patches were incubated in a buffered GA solution for three steps of 24 hours each in a dark room.
  • a GA solution is a 0.6% ⁇ 0.5% v/v for the first and second steps followed by a 0.2% ⁇ 0.15% v/v for the third ones.
  • GA- treated pericardial patches were subjected to two washing steps in phosphate buffer for 15 minutes each. Subsequently, the samples were treated with a polyphenols-based solution under moderate but constant stirring in the dark, for two-step of 25 ⁇ 10 minutes each, at room temperature (RT). At the end of the incubation, the treated patches were subjected to five washes in phosphate buffer for 15 to 30 minutes each.
  • the frames containing the pericardial patch are subjected for a time between 1 and 24 hours to treatment in a hypotonic solution containing an anionic detergent.
  • the treatment is carried out at a temperature between 0°C and 40°C (in the example it was set at 4°C) under moderate but constant stirring.
  • a series of washes with a hypotonic hydroalcoholic solution have been performed.
  • the frames containing the tissue samples move forwards for a time between 1 and 24 hours (in the example described 8 hours) to treatment in an isotonic solution containing a non-anionic detergent followed by a series of washes with an isotonic solution.
  • the frames containing the samples move forward for a time between 1 and 24 hours (in the example described 8 hours) to treatment in an isotonic hydroalcoholic solution.
  • a series of washes with an isotonic solution have been performed.
  • the frames containing the tissue samples are subjected to treatment in an isotonic polyphenols-base buffer.
  • the treatment is carried out at a temperature between 0°C and 40°C (in the example it was set at 20°C) under moderate but constant stirring in the dark.
  • washings in an isotonic solution have been performed.
  • the non-sterile components of the solution are sodium metabisulfite and reduced glutathione, both in powder form. 4% p/v of sodium metabisulfite and 1 % p/v of glutathione were weighed. Subsequently, the proper amount of sterile WFI was added to the powders (20% p/v of the desired final volume). After the addition of WFI, the bottles were placed on an orbital shaker and shaken for 10-15 minutes at room temperature at a speed of 100-200 rpm. Once the powders have been completely dissolved, the resulting liquid appears clear and slightly yellow.
  • Phase 2 Preparation of the sterile solution from the "non-sterile" components
  • Phase 3 Final preparation of the sterile storage solution
  • the final preparation of the sterile solution was carried out under aseptic conditions, working inside an ISO5 Biohazard class biological cabinet.
  • a new sterile bottle was transferred 80% of the final volume of >99% pure glycerol (sterile).
  • the sterile solution prepared in Phases 1 and 2 was added to the glycerol.
  • the container was hermetically sealed and the solution was externally coded as Preserving Solution.
  • the container was removed from the sterile hood and placed on a universal orbital shaker. The solution was stirred slowly for 10-15 minutes at room temperature at a speed of 100-150 rpm to ensure homogeneous distribution of the ingredients.
  • Preserving Solution resulted in transparent and slightly straw yellow.
  • the solution was stored in the refrigerator at +4°C and protected from light.
  • bovine pericardial patches were incubated in the Preserving Solution for up to 36 months. At different time intervals (TO, 15, and 30 days and 2, 4, 8, 16, 24, 32, and 36 months) 4ml of solution was collected with a sterile procedure. The samples were subjected to spectrographic examination (with a wavelength range from 300 nm to 800 nm) with a spectrophotometer (Skyscan, Thermoscientific). For each type of tissue patch, the stability of the Preserving Solution was compared with that of a standard solution. The variation of the spectra over time indicates the occurrence of degradation phenomena affecting the solution itself and the samples.
  • G tissue patch Preserving Solution vs 0.2% v/v buffered glutaraldehyde solution.
  • Figure 2 indicates how all the withdrawals from the Preserving Solution taken at different time intervals return the same type of spectrum with minimal variations. In Figure 3, however, it is possible to highlight how the absorbance values tend to increase constantly over time, suggesting the onset of degradation phenomena affecting the tissue or the solution itself.
  • GP tissue patch Preserving Solution vs isotonic phosphate buffer + 0.04% NaNs.
  • Figure 4 indicates how all the withdrawals from the Preserving Solution taken at different time intervals return the same type of spectrum with minimal variations.
  • Figure 5 the absorbance values tend to increase constantly over time, suggesting the onset of degradation phenomena affecting the tissue or the solution itself.
  • P tissue patch Preserving Solution vs isotonic phosphate buffer + 0.04% NaNs.
  • Figure 6 indicates how all the withdrawals from the Preserving Solution taken at different time intervals return the same type of spectrum with minimal variations.
  • Figure 7 the absorbance values tend to increase constantly over time, suggesting the onset of degradation phenomena affecting the tissue or the solution itself.
  • the Bactericidal Activity (BA) of the Preserving Solution object of the invention was evaluated regarding the following different micro-organism: Staphyloccoccus aureus ATCC 6538, Pseudomonas aeruginosa ATCC 9027, Enterococcus faecalis ATCC 29212, Listeria monocytogenes ATCC 191 1 1 , Salmonella enterica -typhimurium ATCC 14028, Streptococcus viridans ATCC 6249, a nontuberculous mycobacterium Mycobacterium chelonae ATCC 35752, a yeast Candida albicans ATCC 10231 and a fungus Aspergillus brasiliensis ATCC 16404.
  • BA assay consists of a suspension method with a single incubation for 24 hours of the bacteria with a known amount of the Preservative Solution of the invention (inoculum). At the end of the incubation time, the content of each test tube is seeded into 90 mm sterile Petri dishes in a specific agar medium, with a pour plate or spread plate technique depending on the micro-organisms tested, after dilution in Tryptone Salt Broth (MRD Broth). Then the plates obtained are incubated at specific conditions and temperatures according to the growth requirements of each micro-organism.
  • MRD Broth Tryptone Salt Broth
  • a microbial suspension in MRD Broth was quantified through the spectrophotometer at 620 nm wavelength in a disposable 10 mm path length cuvette.
  • the absorbance of an aliquot part of the suspension is measured: the range between 0.150 and 0.460 corresponds to a concentration of cells between 1x10 8 CFU/ml and 3x10 8 CFU/ml (with Candida albicans between 1x10 7 CFU/ml and 3x10 7 CFU/ml).
  • Streptococcus oralis as there was no correlation between the absorbance measure and the bacteria concentration, the quantification was performed by cell count at the microscope.
  • the Preserving Solution object of the invention has proved not to alter the appearance of the tissues contained in it and not to be subject to degrading phenomena affecting the solution itself.
  • the Preserving Solution showed an excellent bactericidal activity comparable to that of a standard antibiotic solution, thus proving effectiveness in preserving the biological material from infections.

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  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
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Abstract

La présente invention concerne une préparation pour le stockage de tissus biologiques.
PCT/IB2024/055741 2024-06-12 2024-06-12 Solution de conservation Pending WO2025257595A1 (fr)

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20100151435A1 (en) * 2007-02-17 2010-06-17 President And Fellows Of Harvard College Compositions and methods for tissue preservation
WO2017093147A1 (fr) 2015-11-30 2017-06-08 Biocompatibility Innovation Srl Procédé d'inactivation de hétéro-antigènes dans des tissus biologiques
US20220145235A1 (en) * 2019-03-15 2022-05-12 Megakaryon Corporation Storage liquid for mammalian cells
WO2020234845A1 (fr) 2019-05-22 2020-11-26 Biocompatibility Innovation Srl Procédé pour empêcher la formation de dépôts calcifiés et pour inactiver des xénoantigènes dans des matrices biologiques
WO2023057947A1 (fr) 2021-10-06 2023-04-13 Biocompatibility Innovation S.r.l. Procédé de préparation de surfaces, en particulier de prothèse cardiaque

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