WO2014201154A1 - Combinaisons de matrices pour permettre la culture in vitro de cellules souches hématopoïétique - Google Patents

Combinaisons de matrices pour permettre la culture in vitro de cellules souches hématopoïétique Download PDF

Info

Publication number
WO2014201154A1
WO2014201154A1 PCT/US2014/041968 US2014041968W WO2014201154A1 WO 2014201154 A1 WO2014201154 A1 WO 2014201154A1 US 2014041968 W US2014041968 W US 2014041968W WO 2014201154 A1 WO2014201154 A1 WO 2014201154A1
Authority
WO
WIPO (PCT)
Prior art keywords
collagen
fibronectin
retronectin
composition
ecm
Prior art date
Application number
PCT/US2014/041968
Other languages
English (en)
Inventor
Marie Zhang
Naira Serobyan
Original Assignee
Microstem, Inc.
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 Microstem, Inc. filed Critical Microstem, Inc.
Publication of WO2014201154A1 publication Critical patent/WO2014201154A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/04—Flat or tray type, drawers
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
    • C12M25/14—Scaffolds; Matrices
    • 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
    • C12N5/0068—General culture methods using substrates
    • 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
    • C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
    • C12N5/0662—Stem cells
    • C12N5/0665—Blood-borne mesenchymal stem cells, e.g. from umbilical cord blood
    • 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
    • C12N2533/00—Supports or coatings for cell culture, characterised by material
    • C12N2533/50—Proteins
    • C12N2533/52—Fibronectin; Laminin
    • 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
    • C12N2533/00—Supports or coatings for cell culture, characterised by material
    • C12N2533/50—Proteins
    • C12N2533/54—Collagen; Gelatin
    • 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
    • C12N2533/00—Supports or coatings for cell culture, characterised by material
    • C12N2533/90—Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
    • 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
    • C12N2535/00—Supports or coatings for cell culture characterised by topography
    • C12N2535/10—Patterned coating

Definitions

  • the present invention relates to methods, compositions and substrates useful for the attachment and proliferation of hematopoietic stem cells.
  • HSCs Hematopoietic stem cells
  • MSCs are the blood cells that give rise to all the blood cell types from the myeloid and lymphoid lineages. HSCs are found in the bone marrow of adults. They are also found in umbilical cord blood and in small numbers in peripheral blood. HSCs are defined by their ability to differentiate to all blood cell types (Multipotency) and their ability to self-renew. Studies have shown that HSCs' self-renewability occurs in the stem cell niche in the bone marrow. HSCs can be expanded in vitro when stimulated using certain combinations of cytokine cocktails. However, it has also been shown that while HSCs can be expanded under such conditions, they also spontaneously differentiate into other more mature blood cell types.
  • HSCs Inside of a body, HSCs reside in a microenvironment (stem cell niche) in bone marrow that determines their self-renewability and differentiation potential.
  • microenvironment consists of extracellular matrices (ECMs), signaling molecules, cytokines, glycans,
  • the ECM as a major microenvironment component provides not only a scaffold for cellular support, but also a storage reservoir for growth factors, hormones and cytokines.
  • HSCs are treated as a non-adherent cell type because of the difficulty in re-creating such microenvironment.
  • a combinatorial ECM component screening device has been designed and manufactured to enable researchers to rapidly identify the appropriate individual components (or combination of components) that constitute an ECM for the optimal culturing of a particular cell type.
  • the invention screening device allows researchers to test up to 10,000 ECM components (or combinations of components) using as little as 20,000 cells.
  • the identified ECM and its combinations enable hematopoietic stem cells to attach and expand as adherent cells on the surface of standard cell culturing devices.
  • the invention device is applicable to all mammalian cell types with a potential for adherence to an ECM or combination of ECMs in a standard tissue culture vessel, including hematopoietic stem cells.
  • Figure 1 illustrates an exemplary ECM array slide design.
  • Each block contains a unique ECM condition with 9 replicate spots.
  • Spot diameter is 400 ⁇ .
  • the distance between spots in each block is 500 ⁇ from center to center.
  • the distance between each block is 4.5 mm.
  • Figure 2 presents an example of ECM conditions on the matrix screen device.
  • Figure 3A illustrates HSC proliferation on an ECM combination of retronectin and fibronectin from 48 to 72 hours.
  • Figure 3B demonstrates that retronectin and fibronectin ECM combination also support CD34 marker expression.
  • FIG. 4 illustrates that retronectin combinations show the most HSC cell attachment after 5 days of culturing. Retronectin alone showed less HSC cell attachment compared to the Retronectin combinations with Fibronectin and with Tropoelastin. Fibronectin alone showed less cell attachment compared to retronectin combinations with Fibronectin and with Tropoelastin and retronectin alone. DETAILED DESCRIPTION OF THE INVENTION
  • compositions comprising:
  • At least one collagen selected from the group consisting of Collagen I, Collagen
  • tropoelastin and/or
  • compositions may further comprise fibronectin.
  • such compositions may further comprise Collagen VI.
  • the preceding compositions may further comprise at least one collagen selected from the group consisting of Collagen I, Collagen III, Collagen IV, Collagen V and Collagen VI (if not already present).
  • the preceding compositions may further comprise Collagen I.
  • the preceding compositions may further comprise Collagen III.
  • the preceding compositions may further comprise Collagen IV.
  • the preceding compositions may further comprise Collagen V.
  • the preceding compositions may further comprise Collagen VI (if not already present).
  • any of the preceding compositions may further comprise laminin.
  • any of the preceding compositions may further comprise vitronectin.
  • any of the preceding compositions may further comprise tropoelastin.
  • any of the preceding compositions may further comprise tenascin.
  • invention compositions comprise retronectin, Collagen I, Collagen III, Collagen IV, Collagen V, Collagen VI, laminin, vitronectin, fibronectin, tropoelastin, and tenascin.
  • the concentration of each of the ECM components, when present, falls in the range of about 0.01 mg/ml up to about 2 mg/ml; in some embodiments, the concentration of each of the ECM components, when present, falls in the range of about 0.02 mg/ml up to about 1.8 mg/ml; in some embodiments, the concentration of each of the ECM components, when present, falls in the range of about 0.05 mg/ml up to about 1.6 mg/ml; in some embodiments, the concentration of each of the ECM components, when present, falls in the range of about 0.1 mg/ml up to about 1.2 mg/ml.
  • compositions comprising:
  • At least one collagen selected from the group consisting of Collagen I, Collagen
  • tropoelastin and/or
  • compositions comprising: Collagen VI, and
  • tropoelastin and/or
  • substrates for attachment of hematopoietic stem cells thereto comprising a support having at least one of retronectin, fibronectin and/or Collagen VI coated thereon.
  • the support is a plate, a slide, a flask, a bead, a chip, and the like.
  • the support comprises a glass slide having 20-100 ⁇ thickness of hydrogel thereon.
  • the support further comprises one or more additional
  • said additional components selected from:
  • At least one collagen selected from the group consisting of Collagen I, Collagen III,
  • tropoelastin and/or
  • compositions comprising a chip comprising a plurality of extracellular matrix (ECM) spots thereon, wherein said ECM comprises:
  • At least one collagen selected from the group consisting of Collagen I, Collagen
  • tropoelastin and/or
  • each ECM spot is 50-1000 ⁇ in diameter
  • center-to-center distance of the nearest spots is sufficient to preclude overlap of the spots.
  • hematopoietic stem cells to a substrate comprising a support having at least
  • the substrate further comprises fibronectin.
  • the substrate further comprises Collagen VI.
  • the culture conditions comprise a temperature of about 37°C and 5% C0 2 .
  • suitable media comprises base media supplemented with cytokines, such as SCF, Flt3, TPO, and the like.
  • cytokines such as SCF, Flt3, TPO, and the like.
  • hematopoietic stem cells to the coated surface (e.g., glass slide or chip) as described herein, and
  • a suitable support e.g., a standard glass slide (75 mm x 25 mm x 1 mm) is coated with a 20-100 ⁇ thickness hydrogel (e.g., polyacrylamide gel) pad.
  • hydrogel e.g., polyacrylamide gel
  • components as well as others, such as growth factors, signaling molecules, antibodies, etc.
  • ECM spots are 50 - 1000 ⁇ in diameter.
  • Each ECM is printed in replicates of 3-20.
  • the center to center distance of nearest spots is sufficient to preclude overlap of the spots (typically the distance between spots is 100 ⁇ or more, preferably >150 ⁇ ). Markings on the glass slide indicate the rows and columns for easy identification of different ECM(s).
  • Figure 1 provides a schematic diagram of an example array design. Each block on the slide is a group of replicates of a particular ECM. An example of ECM conditions designed specifically for attachment and growth of HSCs are listed in Figure 2.
  • the slides are then silanized for 1 hr to overnight in a 2% solution of 3-(trimethoxysilyl) propyl methacrylate in anhydrous toluene, then rinsed in toluene, dried with compressed air, and baked for 15 min to 1 hour in an oven (65°C).
  • a 40- 100 ⁇ . of solution of 10.5% (w/v) acrylamide, 0.55% (w/v) bisacrylamide, 10% (w/v) photoinitiator Irgacure 2959, Ciba Specialty Chemicals 12959 (200 ⁇ / ⁇ ⁇ , in 100% methanol) is placed on a silanized slide and covered with a 75 mm x25 mm or 65 mm x 25 mm cover slip.
  • the slide is then exposed to 1.5 mW/cm 2 365-nm ultraviolet A light for 10 min and immersed in MQH 2 0 for 2 min.
  • the cover slip is then removed, leaving a thin (40-80 ⁇ thickness) polyacrylamide gel pad.
  • the polyacrylamide slides are soaked in MQH 2 0 overnight, and then dried on a hot plate (40°C) for 10 min (See Figures 3 A and 3B).
  • ECM components (0.01 mg/ml - 1 mg/ml), such as, but not limited to Collagen I, III, IV, V, VI, Fibronectin, Laminin, Vitronectin, TropoElastin, Tenascin, Retronectin, Poly-D-lysine as well as CD34 antibody are mixed 1 : 1 with 200 mM acetate, 10 mM EDTA, 40% (v/v) glycerol, and 0.5% (v/v) Triton X-100 in MQH 2 0, at pH 4.9-8.5.
  • ECM components such as, but not limited to Collagen I, III, IV, V, VI, Fibronectin, Laminin, Vitronectin, TropoElastin, Tenascin, Retronectin, Poly-D-lysine as well as CD34 antibody are mixed 1 : 1 with 200 mM acetate, 10 mM EDTA, 40% (v/v) glycerol, and 0.5% (v/v
  • a sterile container such as Nunc rectangle culture plate (Thermo Fisher) soaked in PBS while being exposed to UVC germicidal radiation in a sterile flow hood for minimum 30 min.
  • Frozen CD34+ cells from cord blood were thawed in 37°C water bath and resuspended in X-vivo 20 (Lonza) or StemSpan (Stem Cell Technologies) media supplemented with cytokines, SCF, Flt3 and TPO (10-50ng/ml each). 5 ml of cell suspension (total 50-70,000 cells) was added to the device that was placed in a 4 well NUNC cell culture vessel.
  • the cells were allowed to attach for 2-5 days in an incubator at 37°C and 5% C0 2 .
  • the vessel containing the device was removed from the 37 C incubator. Media was removed and new media with the same cytokine supplement were added. Cells on the device were observed under a phase contrast microscope. Cell attachments on some of the spot conditions were observed. Cells were cultured for one additional day (total 3 days). Media was aspirated from the vessel containing the device. Device were then washed with PBS twice and 4% paraformaldehyde (PFA) made in PBS was added to the vessel containing the device for 15 min at room temperature to fix the cells. PFA was aspirated and the device was washed with PBS. Cells were then stained with CD34 antibody. The slide was imaged using a Leica fluorescence microscope. Results are presented in Figures 3 A and 3B.
  • CD34+ cell from cord blood cell were allowed to culture on the device for a total of 5 days. Significant attachments were observed in many of the spots, see Figure 4.
  • the ECM combination of retronectin and fibronectin was then coated on a 96 well plate.
  • CD34+ cord blood cells were cultured in the presence of StemSpan media supplemented with SCF, Flt3 and TPO. Greater than 90% of the cells were attached onto the retronectin- and fibronectin-coated surface. Small molecule library screens were performed on these adherent CD34+ cells from cord blood to look for factors to further enhance proliferation while maintaining CD34 marker expression.
  • Retronectin in combination with fibronectin, laminin, vitronectin, tropoelastin and tenascin shows significant HSC attachment; the retronectin/fribonectin combination has the most cell attachment.
  • Retronectin in combination with collagen I, III, IV, V and VI, as well as with fibronectin+vitronectin and vitronectin+laminin shows visible but less HSC attachment.
  • cells on retronectin show the most attachment, Fibronectin shows less and Collagen VI shows the least amount of cell attachment.
  • Other ECMs such as collagen I, III, IV, V, laminin, vitronectin, tropoelastin and tenascin alone do not show HSC attachments.
  • CD 34 expression analysis shows that many retronectin ECM combinations support a certain degree of CD34 expression (stemness) with the retronectin and fibronectin combination supporting the highest degree of CD34 expression (>90% of cells are CD34+).

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Genetics & Genomics (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • General Engineering & Computer Science (AREA)
  • Developmental Biology & Embryology (AREA)
  • Cell Biology (AREA)
  • Sustainable Development (AREA)
  • Biophysics (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicinal Chemistry (AREA)
  • Rheumatology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Toxicology (AREA)
  • Immunology (AREA)
  • Hematology (AREA)
  • Clinical Laboratory Science (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

La présente invention concerne un dispositif de criblage de composants de MEC combinatoires conçu et fabriqué pour permettre aux chercheurs d'identifier rapidement les composants individuels ou les combinaisons de composants appropriés qui constituent une MEC pour la culture optimale d'un type de cellules particulier. Le dispositif de criblage de l'invention permet aux chercheurs de tester jusqu'à 10 000 composants de MEC (ou combinaisons de composants) en utilisant seulement 20 000 cellules. La MEC identifiée et ses combinaisons permettent l'attachement des cellules souches hématopoïétiques et leur expansion sous forme de cellules adhérentes sur la surface de dispositifs de culture de cellules standard. Le dispositif de l'invention est applicable à tous les types de cellules de mammifères présentant un potentiel d'adhérence à une MEC ou une combinaison de MEC dans un récipient de culture de tissu standard, notamment les cellules souches hématopoïétiques.
PCT/US2014/041968 2013-06-14 2014-06-11 Combinaisons de matrices pour permettre la culture in vitro de cellules souches hématopoïétique WO2014201154A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US201361835394P 2013-06-14 2013-06-14
US61/835,394 2013-06-14

Publications (1)

Publication Number Publication Date
WO2014201154A1 true WO2014201154A1 (fr) 2014-12-18

Family

ID=52022746

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2014/041968 WO2014201154A1 (fr) 2013-06-14 2014-06-11 Combinaisons de matrices pour permettre la culture in vitro de cellules souches hématopoïétique

Country Status (1)

Country Link
WO (1) WO2014201154A1 (fr)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006078814A2 (fr) * 2005-01-20 2006-07-27 The Regents Of The University Of California Microreseaux cellulaires pour le criblage de facteurs de differenciation
US20100021998A1 (en) * 2008-07-25 2010-01-28 Becton, Dickinson And Company Defined cell culturing surfaces and methods of use
US20120009676A1 (en) * 2010-06-15 2012-01-12 Amanda Mack Generation of induced pluripotent stem cells from small volumes of peripheral blood

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006078814A2 (fr) * 2005-01-20 2006-07-27 The Regents Of The University Of California Microreseaux cellulaires pour le criblage de facteurs de differenciation
US20100021998A1 (en) * 2008-07-25 2010-01-28 Becton, Dickinson And Company Defined cell culturing surfaces and methods of use
US20120009676A1 (en) * 2010-06-15 2012-01-12 Amanda Mack Generation of induced pluripotent stem cells from small volumes of peripheral blood

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
FENG, QI ET AL.: "Expansion of engrafting human hematopoietic stem/ progenitor cells in three-dimensional scaffolds with surface-immobilized fibronectin", JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A, vol. 78, no. 4, 2006, pages 781 - 791, XP002602539, DOI: doi:10.1002/JBM.A.30829 *
TIWARI, ABHILASHA ET AL.: "Ex vivo expansion of haematopoietic stem/ progenitor cells from human umbilical cord blood on acellular scaffolds prepared from MS-5 stromal cell line", JOURNAL OF TISSUE ENGINEERING AND REGENERATIVE DEDICINE, vol. 7, no. 11, 17 April 2012 (2012-04-17), pages 871 - 883 *

Similar Documents

Publication Publication Date Title
De Silva et al. Micro-patterning of animal cells on PDMS substrates in the presence of serum without use of adhesion inhibitors
CN101842474A (zh) 表面柔软的多孔培养平板
Witkowska-Zimny et al. Effect of substrate stiffness on differentiation of umbilical cord stem cells
US8993322B2 (en) Methods and kits for cell release
US20100304427A1 (en) Substrates for adhering, culturing and assaying cells
JP2021158960A (ja) 多能性幹細胞から外胚葉系細胞を分化誘導する方法及び製造方法
JP2008173018A (ja) 細胞培養方法及び細胞培養用基板
CN113846016B (zh) 一种高通量多孔阵列芯片、装置、制备方法及应用
WO2014201154A1 (fr) Combinaisons de matrices pour permettre la culture in vitro de cellules souches hématopoïétique
JP2016063801A (ja) 細胞培養用支持体と、それを用いた細胞培養方法
TW202242093A (zh) 細胞培養基質、其方法及用途
WO2012137830A1 (fr) Kit de culture de cellules animales, procédé de culture de cellules animales, procédé de culture sélective de cellules animales et procédé de différentiation cellulaire
JP2755880B2 (ja) 培養用器具及びその製造方法
JP6145324B2 (ja) 神経細胞への分化誘導を促進させる細胞培養基材
EP3540041A1 (fr) Procédé, appareil et programme de production de tissu cellulaire
US20200332252A1 (en) Hollow microcarrier for shear-free culture of adherent cells in bioreactors
EP3885432A1 (fr) Procédé de fabrication d'un substrat sur lequel sont disposées des cellules nerveuses
JP2022002504A (ja) 神経細胞が配置された基板の製造方法
JP2012520667A (ja) 細胞生育用のポリマー
Nie et al. Polydopamine-mediated surface modification promotes the adhesion and proliferation of human induced pluripotent stem cells
WO2023027079A1 (fr) Dispositif de production de structure cellulaire
WO2022230734A1 (fr) Substrat de culture cellulaire et son procédé de production, procédé d'induction de la différenciation d'une cellule souche pluripotente, et kit de culture cellulaire
JP6332646B2 (ja) 活性エステル基を含有するシラン化合物とそれを用いた材料
US20240218322A1 (en) Cell culture substrate and method for producing same, method for inducing differentiation of pluripotent stem cell, and cell culture kit
WO2023282253A1 (fr) Matériau de base de culture cellulaire pour milieu sans sérum

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 14810191

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

32PN Ep: public notification in the ep bulletin as address of the adressee cannot be established

Free format text: NOTING OF LOSS OF RIGHTS PURSUANT TO RULE 112(1) EPC (EPO FORM 1205A DATED 13.04.2016)

122 Ep: pct application non-entry in european phase

Ref document number: 14810191

Country of ref document: EP

Kind code of ref document: A1