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WO2008108549A1 - Procédé de conservation structurelle d'hémocytes à long terme reposant sur une technique de lyophilisation cellulaire - Google Patents

Procédé de conservation structurelle d'hémocytes à long terme reposant sur une technique de lyophilisation cellulaire Download PDF

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Publication number
WO2008108549A1
WO2008108549A1 PCT/KR2008/001119 KR2008001119W WO2008108549A1 WO 2008108549 A1 WO2008108549 A1 WO 2008108549A1 KR 2008001119 W KR2008001119 W KR 2008001119W WO 2008108549 A1 WO2008108549 A1 WO 2008108549A1
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Prior art keywords
concentration
hemocytes
cell
trehalose
hours
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PCT/KR2008/001119
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English (en)
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Jootae Kim
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Priority claimed from KR1020070021350A external-priority patent/KR100868602B1/ko
Priority claimed from KR1020070023828A external-priority patent/KR20080083376A/ko
Application filed by Individual filed Critical Individual
Publication of WO2008108549A1 publication Critical patent/WO2008108549A1/fr
Anticipated expiration legal-status Critical
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0634—Cells from the blood or the immune system
    • C12N5/0641—Erythrocytes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2500/00—Specific components of cell culture medium
    • C12N2500/30—Organic components
    • C12N2500/32—Amino acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2500/00—Specific components of cell culture medium
    • C12N2500/30—Organic components
    • C12N2500/36—Lipids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2500/00—Specific components of cell culture medium
    • C12N2500/30—Organic components
    • C12N2500/38—Vitamins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2501/00—Active agents used in cell culture processes, e.g. differentation
    • C12N2501/30—Hormones
    • C12N2501/33—Insulin
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2501/00—Active agents used in cell culture processes, e.g. differentation
    • C12N2501/80—Neurotransmitters; Neurohormones
    • C12N2501/825—Serotonine (5-HT); Melatonine

Definitions

  • the field of this invention belongs under cell preservation using lyophilization technique. If this method is utilized for preservation of eukaryotic cell, lyophilized cell can be stabilized structurally and can be preserved without cell damage over a long period of time.
  • Method related with this study is to load high treahlose level in the extracellular space and then to increases trehalose concentration of cell inside.
  • This method has advantage that cell membrane is not manipulated artificially. But there is a problem that cell damage increases because of high treahalose level in itself. If erythrocjtes are incubated during 24 hours, the half degree of the cells become hemolytic. This hemolytic erythrocyte should be removed one by one to use clinically, but this technique is very difficult. Moreover non-hemolytic erythrocyte can not be regarded healthy for its weakness. So using this method, technique that stabilze the cell and cell membrane should be developed.
  • Vacuum condition can do protection of microoiganisms by minimizing oxidation, but there are a problems in case that preserve eukaryotic cell; it is impossible to make high vacuum condition that gas does not exist totally, the minimum gas induce eukaryotic cell to cell oxidation because eukaryotic cell don't have genetic mechanism that stoping cellular function in the dehydrated state. This oxidation produce free radical, it destroys cell oiganelle and cell membrane. Method that solve this problem is to find the stable materials for preserving eukaryotic cell. Disclosure of Invention Technical Problem
  • the first subject that should solve is as following. When cells are loaded with trehalose of high concentration, cell damage should be reduced to clinical usable level. To use clinically, next two conditions must be satisfied; First, The rate of cell damage must become around 5%. Second, trehalose concentration of cell inside must become more than 6OmM. It is not difficult to make cell model loaded to high trehalose level. Simply trehalose and antioxidant have only to be put on the incubated cells. But it is very difficult that determinate concentration and preparation of trehalose and antioxidant. Several kinds of antioxidant must be put on one by one and observe result one after another. Previous our study revealed that insulin increased antioxidant function on hemolysis of red blood cell. So, mixture of antioxidant and insulin were loaded on the same cell model.
  • glycerol is selected among several materials.
  • Third, water absorptiveness of glycerol is very high, but if there does not exits water in storage receptacle, there will be no circumstance to absorb water. And glycerol exits liquid state in room temperature. Fourth, glycerol is very low toxicity and low cell permeability. Fifth, glycerol has no explosiveness, and there is no any change of state in room temperature.
  • the model of cells to preserve is human erythrocytes.
  • the antioxidants are selected all materials including with ⁇ -tocopherol, ⁇ -lipoic acid, melatonin, N- acetyl-L-cysteine, L-ascorbic acid which is good effect from previous hemolysis experiment. Insulin is used as material which rises an effect of antioxidant.
  • PBS Phosphate-buffered saline, 300mOsm, pH 7.2
  • ADSOL (4ffim ⁇ sm) contained 11 ImM glucose, 2mM adenine, 154mM NaCl and 4ImM mannitol.
  • Buffer PBS(T) contained PBS and indicated amount of trehalose.
  • Buffer ADSOL(T) contained ADSOL and K-phosphate and indicated amount of trehalose.
  • ADSOL(T) (80OmM) contained 80OmM endotoxin- free trehalose (Hayashibara biochemical lab.
  • ⁇ - tocopherol was prepared as aqueous solution including l%(v/v) ethanol.
  • ⁇ -lipoic acid and melatonin was prepared as aqueous solution including 0.5%(v/v) ethanol. This use of ethanol did not show no significant change in previous study of red blood cell hemolysis. Order of experiments, measuring methods, results of experiment were as followings
  • erythrocytes were collected from the bottom portion of the packed red cells. Then the cells were washed in PBS and centrifuged at 515g, lOmin three times. At each spin, erythrocytes were collected same method. Finally the erythrocytes were stored in PBS at 4 0 C with 30% hematocrit and used within 1 day.
  • the loaded cells (0.5ml 30% hematocrit) were centrifugated at 350g for 3min and were mixed with 4ml of 80% methanol. The mixture was incubated at 85 0 C for 45min followed by centrifugation at 20Og for lOmin. The supernatant was collected and evaporated using a vacuum centrifuge(Labconco, USA), and dry residue was dissolved in 3mL of nano-pure water. This sample (3mL) were mixed with 6mL of anthrone reagent [2% anthrone in concentrated sulfuric acid], heated to 100 0 C for 3min, and allowed to cool.
  • anthrone reagent 2% anthrone in concentrated sulfuric acid
  • Absorbance at 620nm was measured on a spectrophotometer (Hitachi U2000, Japan)at room temperature and compared with a standard curve. Since the anthrone method detects all sugars, unloaded control erythrocytes were always treated in parallel. These values, normalized for cell count, were subtracted from the trehalose specifically and to avoid artifact due to endogenous sugars. Data are shown for at least three independent experiments.
  • Figure 1 presents the result of incubated erythrocytes in 80OmM trehalose buffer without cell protection protocol for 15 hours in 37 0 C.
  • erythrocytes were incubated during 15 hours ( ⁇ +4. ImM)
  • this result showed that trehalose concentration of cell interior becomed above 6OmM.
  • Figure 2 presents degree of erythrocytes hemolysis in 80OmM trehalose buffer without cell protection protocol in 37 0 C.
  • erythrocytes were incubated for 15 hours(25 ⁇ 3.1%), erythrocytes showed 25% hemolysis.
  • This result showed that erythrocytes were available to use clinically if only hemolytic erythrocytes were removed.
  • this result showed that 5% hemolysis happened when 3 hours passed. Therefore, cell protection protocol must decrease cell damage up to result of 3 hours hemolysis statistically.
  • Figure 3 presents the degree of hemolysis under each conditions. A(5.2+0.69%) and
  • Figure 4 presents the result of incubated erythrocytes in 80OmM trehalose buffer with the final cell protection protocol ( ⁇ - tocopherol, ⁇ - lipoic acid, melatonin, N- acetyl-L-cysteine, L-ascorbic acid, insulin) in 37 0 C. It showed that concentration of cell inside beconed above 6OmM when erythrocytes were incubated for 15 hours (68+3.ImM). This value showed that there was no statistical significance to compared to value of result which was incubated for 15 hours (69+4.ImM) in Figure 1.
  • the model of cells to preserve is human hemocytes.
  • the names and concentration of cell protection material are 5 x 10 '4 M L-ascorbic acid, 5 x 10 '5 M ⁇ -lipoic acid, 5 x 10 ⁇ 5 M Melatonin, 3 x 10 4 M N-acetyl-L-cysteine, 10 5 M ⁇ -tocopherol, and 5 x 10 5 M insulin.
  • the 10 5 M prostaglandin El is also used because prostaglandin El did role of stabilizing platelet in our previous experiment.
  • PBS Phosphate-buffered saline, 300mOsm, pH 7.2
  • ADSOL 462mOsm
  • ADSOL contained 11 ImM glucose, 2mM adenine, 154mM NaCl and 4ImM mannitol.
  • Buffer PBS(T) contained PBS and indicated amount of trehalose.
  • Buffer ADSOL(T) contained ADSOL and K-phosphate and indicated amount of trehalose.
  • ADSOL(T) (80OmM) contained 80OmM endotoxin-free trehalose (Hayashibara biochemical lab.
  • Lyophilizing buffer contained lOOmOsmol ADSOL(24mM glucose, 0.43mM adenine, 33mM NaCl, 89mM mannitol), 10OmM trehalose, 66mM K-phosphate, 15% high molecular weight hydroxyethyl starch (HES, B. Braun Medical, Irvine, CA) and 2.5% human serum albumin (HSA, Sigma- Aldrich).
  • ⁇ -tocopherol was prepared as aqueous solution including l%(v/v) ethanol.
  • ⁇ -lipoic acid, melatonin and prostaglandin were prepared as aqueous solution including 0.5%(v/v) ethanol.
  • erythrocytes were collected from the bottom portion of the packed red cells, leukocytes were collected from the buffy coat, and platelets were collected from upper layer of sample. Each hemocytes were collected to same tube. Then the cells were washed in PBS and centrifuged at 515g, lOmin three times. At each spin, blood corpuscles were collected same method. Washed erythrocytes were stored in PBS at 4 0 C with 30% hematocrit and used within 1 day.
  • lyophilizer (Labconco,USA) The conditions of lyophilizer (Labconco,USA) were set as following. Hemocytes were incubated for 15 hours and were added with lyophilization buffer until the hematocrit reached to 5%. Then, the hemocytes were cooled at a rate of I 0 C per minute until they got to -4O 0 C, in which temperature the hemocytes were incubated for 30 minutes. Primary drying were performed at -3O 0 C for 7 hours at a vacuum of 30mTorr. During secondary drying which were performed for 6 hours, shelf temperature were increased from -3O 0 C to 2O 0 C at a rate of 0.8 0 C per minute using a vacuum of SOmTorr. After then, samples were put into a specially manufactured container. The air in the container were removed and the container were filled with anhydrous glycerol(>99.5%) for a long-term storage.
  • Fig 5 presents well-preserved erythrocytes generally as time passes with little damage of erythrocytes. For the 6 months, cell morphology was showed most well- preserved aspect. Also, because there is no living things that live in the anhydrous glycerol(>99.5%), if basic aseptic method is performed, the contamination to mi- crooiganism, fungus, virus will be not concerned about. However, there are some problem that totally recover function. This problem is next subject that must solve. [41] Fig 6 presents sample that be utilized for remembering deceased people in funeral.
  • the decoration container have lyophilized erythrocytes of the deceased. Even if all of deceased disappears with death, his blood corpuscle cells will be kept over a long period of time. After this is utilized for funeral ceremony, keeping can be with family or deceased.
  • Fig 7 presents sample that be utilized for commemoration between lovers.
  • the decoration container is personal ornaments which is portable commemoration material of lover.
  • Figure 1 shows a graph illustraing the change of trehalose level at intervals of 3 hours when to erythrocytes were added with 80OmM trehalose in 37 0 C.
  • Figure 2 shows a graph illustraing the degree of hemolysis at intervals of 3 hours when to erythrocytes were added with 80OmM trehalose in 37 0 C.
  • Figure 3 shows a graph illustrating the degree of hemolysis occurred in 15 hours observation depending on each situation of cell protection protocols when to erythrocytes were added with 80OmM trehalose in 37 0 C.
  • Figure 4 shows a graph illustrating the change of trehalose level at intervals of 3 hours observation depending on final cell protection protocols when to erythrocytes were added with 80OmM trehalose in 37 0 C.
  • Figure 5 shows the result of long-term storage of lyophilized cells observed depending on time under the phase contrast microscope.
  • Figure 6 shows a sample of lyophilized cells stored using this technique which can be utilized for rememberig deceased people.
  • Figure 7 shows a sample of lyophilized cells stored using this technique which can be utilized for commemoration between lovers.
  • T3 3 hours, T ⁇ 6 hours, T9: 9 hours, T12 12 hours, T15: 15 hours
  • C The condition that erythrocytes were added with 80OmM trehalose, 5 x 10 4 M L- ascorbic acid for 15 hours in 37 0 C.
  • D The condition that erythrocytes were added with 80OmM trehalose, 5 x 10 5 M ⁇ - lipoic acid for 15 hours in 37 0 C.
  • E The condition that erythrocytes were added with 80OmM trehalose, 5 x 10 5 M
  • F The condition that erythrocytes were added with 80OmM trehalose, 3 x 10 4 M N- acetyl-L-cysteine for 15 hours in 37 0 C.
  • G The condition that erythrocytes were added with 80OmM trehalose, 10 5 M ⁇ - tocopherol for 15 hours.
  • H The condition that erythrocytes were added with 80OmM trehalose, 5 x 10 4 M L- ascorbic acid, 5 x 10 5 M ⁇ -lipoic acid, 5 x 10 5 M Melatonin, 3 x 10 4 M N- acetyl-L-cysteine, 10 '5 M ⁇ -tocopherol for 15 hours in 37 0 C.
  • the lyophilized cell can be preserved for remembering deceased people in funeral.
  • the lyophilized cell can be preserved for remembering deceased people in funeral.
  • the lyophilized cell can be preserved for commemoration between lovers.

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  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
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  • Genetics & Genomics (AREA)
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  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

L'invention concerne un procédé permettant de conserver des cellules de manière structurelle pendant une période courte ou longue. Ce procédé repose sur une technique de lyophilisation. Le procédé de l'état de la technique permet de préserver des cellules humaines lyophilisées pendant longtemps mais il présente des limites. Notre institut de recherches a découvert une manière de conserver les cellules lyophilisées de manière structurelle et sûre pendant une période courte ou longue. Le modèle de cellule à conserver est l'hémocyte humain. Il existe deux catégories de procédé de stockage : un procédé de stockage à court terme et un procédé de stockage à long terme. Pour conserver les hémocytes humains à long terme, on fait incuber les hémocytes dans une solution tampon de tréhalose 800nM à 37°C pendant 15 heures. Les noms et la concentration de la matière de protection cellulaire sont: acide L-ascorbique 5 × 10-4M; acide α-lipoïque 5 × 10-5M, mélatonine 5 × 10-5M; N-acétyl-L-cystéine3 × 10-4M; δ-tocophérol 10-5M; insuline 5 × 10-5M et prostaglandine E1 10-5M. Le stockage à long terme commence par la lyophilisation d'hémocytes incubés auquels on ajoute du glycérol plus tard. Les hémocytes incubent pendant 15 heures et sont ajoutés au tampon de lyophilisation jusqu'à ce que l'hématocrite atteigne 5 %. Puis, les hémocytes sont refroidis à une vitesse de 1°C par minute jusqu'à ce qu'ils atteignent -40°C, température à laquelle ils incubent pendant 30 minutes. Un séchage primaire s'effectue à -30°C pendant 7 h sous un vide de 30mTorr. Pendant le séchage secondaire qui dure six heures, la température de conservation augmente de -30 °C à 20 °C à une vitesse de 0, 8°C par minute, avec un vide de 50mTorr. Enfin, des échantillons sont placés dans un contenant spécialement conçu à cet effet. L'air du contenant est retiré et le contenant est rempli de glycérol anhydre (≥99,5%) pour un stockage à long terme. Même si de nombreuses études sont nécessaires pour retrouver les caractéristiques fonctionnelles de manière parfaite, le procédé de l'invention permet d'obtenir une stabilisation structurelle permettant un stockage semi-permanent. Le procédé de l'invention, destiné à stocker des cellules lyophilisées de manière semi-permanente, peut être utilisé de plusieurs façons dans nos vies quotidiennes. Par exemple, il peut être utilisé pour se souvenir de personnes décédées, à des funérailles, ou pour une commémoration entre des amoureux.
PCT/KR2008/001119 2007-03-05 2008-02-27 Procédé de conservation structurelle d'hémocytes à long terme reposant sur une technique de lyophilisation cellulaire Ceased WO2008108549A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
KR1020070021350A KR100868602B1 (ko) 2007-03-05 2007-03-05 인간 적혈구가 고농도 트리할로즈에 노출되었을 때발생하는 세포 용혈 현상을 최소화하는 방법
KR10-2007-0021350 2007-03-05
KR10-2007-0023828 2007-03-12
KR1020070023828A KR20080083376A (ko) 2007-03-12 2007-03-12 세포의 동결건조 기술을 이용해 인간의 혈구세포를구조적으로 장기간 보관하는 방법

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WO2008108549A1 true WO2008108549A1 (fr) 2008-09-12

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Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013087936A1 (fr) * 2011-12-16 2013-06-20 Galderma Research & Development Association d'un agoniste des récepteurs des prostaglandines et d'un agoniste du récepteur mc1r pour le traitement et/ou la prévention de désordres de la pigmentation
WO2015191633A1 (fr) * 2014-06-10 2015-12-17 Biomatrica, Inc. Stabilisation de polypeptides non dénaturés, d'acides nucléiques, et d'exosomes dans un échantillon de sang à des températures ambiantes
US9376709B2 (en) 2010-07-26 2016-06-28 Biomatrica, Inc. Compositions for stabilizing DNA and RNA in blood and other biological samples during shipping and storage at ambient temperatures
US9725703B2 (en) 2012-12-20 2017-08-08 Biomatrica, Inc. Formulations and methods for stabilizing PCR reagents
US9845489B2 (en) 2010-07-26 2017-12-19 Biomatrica, Inc. Compositions for stabilizing DNA, RNA and proteins in saliva and other biological samples during shipping and storage at ambient temperatures
US10064404B2 (en) 2014-06-10 2018-09-04 Biomatrica, Inc. Stabilization of thrombocytes at ambient temperatures
US10568317B2 (en) 2015-12-08 2020-02-25 Biomatrica, Inc. Reduction of erythrocyte sedimentation rate
US11604026B2 (en) 2019-03-14 2023-03-14 Terumo Bct Biotechnologies, Llc Lyophilization loading tray assembly and system
US11634257B2 (en) 2017-10-09 2023-04-25 Terumo Bct Biotechnologies, Llc Lyophilization container and method of using same

Citations (3)

* Cited by examiner, † Cited by third party
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US5242792A (en) * 1991-02-25 1993-09-07 The United States Of America As Represented By The Secretary Of The Navy Method for the preservation of red blood cells by lyophilization using glycerol or inositol with disaccharides
US5965438A (en) * 1995-06-07 1999-10-12 Phyton, Inc. Cryopreservation of plant cells
US20040067480A1 (en) * 2002-10-04 2004-04-08 Organ Recovery Systems, Inc. Method for treatment of cellular materials with sugars prior to preservation

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5242792A (en) * 1991-02-25 1993-09-07 The United States Of America As Represented By The Secretary Of The Navy Method for the preservation of red blood cells by lyophilization using glycerol or inositol with disaccharides
US5965438A (en) * 1995-06-07 1999-10-12 Phyton, Inc. Cryopreservation of plant cells
US20040067480A1 (en) * 2002-10-04 2004-04-08 Organ Recovery Systems, Inc. Method for treatment of cellular materials with sugars prior to preservation

Cited By (22)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9999217B2 (en) 2010-07-26 2018-06-19 Biomatrica, Inc. Compositions for stabilizing DNA, RNA, and proteins in blood and other biological samples during shipping and storage at ambient temperatures
US9376709B2 (en) 2010-07-26 2016-06-28 Biomatrica, Inc. Compositions for stabilizing DNA and RNA in blood and other biological samples during shipping and storage at ambient temperatures
US9845489B2 (en) 2010-07-26 2017-12-19 Biomatrica, Inc. Compositions for stabilizing DNA, RNA and proteins in saliva and other biological samples during shipping and storage at ambient temperatures
WO2013087936A1 (fr) * 2011-12-16 2013-06-20 Galderma Research & Development Association d'un agoniste des récepteurs des prostaglandines et d'un agoniste du récepteur mc1r pour le traitement et/ou la prévention de désordres de la pigmentation
US9364455B2 (en) 2011-12-16 2016-06-14 Galderma Research & Development Combination of a prostaglandin receptor agonist and an MC1R receptor agonist for the treatment and/or prevention of pigmentation disorders
US9725703B2 (en) 2012-12-20 2017-08-08 Biomatrica, Inc. Formulations and methods for stabilizing PCR reagents
US11672247B2 (en) 2014-06-10 2023-06-13 Biomatrica, Inc. Stabilization of thrombocytes at ambient temperatures
WO2015191633A1 (fr) * 2014-06-10 2015-12-17 Biomatrica, Inc. Stabilisation de polypeptides non dénaturés, d'acides nucléiques, et d'exosomes dans un échantillon de sang à des températures ambiantes
US10772319B2 (en) 2014-06-10 2020-09-15 Biomatrica, Inc. Stabilization of thrombocytes at ambient temperatures
US12121022B2 (en) 2014-06-10 2024-10-22 Biomatrica, Inc. Stabilization of thrombocytes at ambient temperatures
US10064404B2 (en) 2014-06-10 2018-09-04 Biomatrica, Inc. Stabilization of thrombocytes at ambient temperatures
US10568317B2 (en) 2015-12-08 2020-02-25 Biomatrica, Inc. Reduction of erythrocyte sedimentation rate
US11116205B2 (en) 2015-12-08 2021-09-14 Biomatrica, Inc. Reduction of erythrocyte sedimentation rate
US12089588B2 (en) 2015-12-08 2024-09-17 Biomatrica, Inc. Reduction of erythrocyte sedimentation rate
US11634257B2 (en) 2017-10-09 2023-04-25 Terumo Bct Biotechnologies, Llc Lyophilization container and method of using same
US11609042B2 (en) 2019-03-14 2023-03-21 Terumo Bct Biotechnologies, Llc Multi-part lyophilization container and method of use
US11740019B2 (en) 2019-03-14 2023-08-29 Terumo Bct Biotechnologies, Llc Lyophilization loading tray assembly and system
US11747082B2 (en) 2019-03-14 2023-09-05 Terumo Bct Biotechnologies, Llc Multi-part lyophilization container and method of use
US11815311B2 (en) 2019-03-14 2023-11-14 Terumo Bct Biotechnologies, Llc Lyophilization container fill fixture, system and method of use
US11994343B2 (en) 2019-03-14 2024-05-28 Terumo Bct Biotechnologies, Llc Multi-part lyophilization container and method of use
US11609043B2 (en) 2019-03-14 2023-03-21 Terumo Bct Biotechnologies, Llc Lyophilization container fill fixture, system and method of use
US11604026B2 (en) 2019-03-14 2023-03-14 Terumo Bct Biotechnologies, Llc Lyophilization loading tray assembly and system

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