IMR90-4 and DF19-9-11T iPSCs and H9 hESCs were maintained between passages 26–42 on Matrigel (BD Biosciences) in mTeSR1 medium (STEMCELL Technologies) or on irradiated mouse embryonic fibroblasts (MEFs) in standard unconditioned medium (UM) as previously described18 (link). For differentiation, cells were passaged onto Matrigel in mTeSR1 medium for 2–3 days of expansion and then switched to unconditioned medium (UM) lacking bFGF for 6 days. Human endothelial serum-free medium (hESFM; Life Technologies) supplemented with 20 ng/mL bFGF (R&D Systems) and 1% platelet-poor plasma derived bovine serum (Biomedical Technologies, Inc.) was then added for an additional 2–4 days. All-trans RA (Sigma) was reconstituted in DMSO and included at concentrations of 1–10 μM depending on the experiment. Cells were then dissociated with Versene (Life Technologies) and plated onto 12-well tissue culture polystyrene plates or 1.12 cm2 Transwell-Clear® permeable inserts (0.4 μm pore size) coated with a mixture of collagen IV (400 μg/mL; Sigma) and fibronectin (100 μg/mL; Sigma) in H2O. Culture plates were incubated with the coating for at least 30 min at 37°C, while the inserts were incubated for a minimum of 4 h at 37°C. Resultant, purified hPSC-derived BMECs were then grown in EC medium for 24 h (with or without RA); in some experiments, primary pericytes or fibroblasts were co-cultured with BMECs during these 24 h (see description below). After this 24 h period, BMECs were continued as monoculture or co-cultured as described below. In our previous publication, we had utilized dispase for subculturing the BMECs18 (link), but non-enzymatic treatment of the BMECs with Versene results in less debris attached to the BMEC monolayer. We had also used hPSCs exclusively maintained on MEFs prior to differentiation18 (link), but in this study no noticeable differences in BBB properties were observed between hPSCs maintained on MEFs and hPSCs maintained under feeder-independent conditions and we now exclusively use hPSCs maintained in mTeSR1 on Matrigel.
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Collagen Type IV
Collagen Type IV
Collagen Type IV is a key structural component of basement membranes, which are specialized extracellular matrices that provide support and organization to various tissues throughout the body.
This collagen type is essential for the integrity and functioning of organ systems, including the kidneys, lungs, and blood vessels.
Reasearchers studying Collagen Type IV can leverage PubCompare.ai's AI-powered platform to optimaize their research protocols and enhance reproducibility.
The platform can effortlessly locate the best protocols from literature, pre-prints, and patents, while providing AI-driven comparisons to identify the most effective products.
This can help take the guesswork out of research and elevate findings on this important collagen type.
This collagen type is essential for the integrity and functioning of organ systems, including the kidneys, lungs, and blood vessels.
Reasearchers studying Collagen Type IV can leverage PubCompare.ai's AI-powered platform to optimaize their research protocols and enhance reproducibility.
The platform can effortlessly locate the best protocols from literature, pre-prints, and patents, while providing AI-driven comparisons to identify the most effective products.
This can help take the guesswork out of research and elevate findings on this important collagen type.
Most cited protocols related to «Collagen Type IV»
Biomedical Technology
Blood Platelets
Bos taurus
Cells
Collagen Type IV
dispase
Embryo
Endothelium
Enzymes
Fibroblasts
FN1 protein, human
Homo sapiens
Human Embryonic Stem Cells
Induced Pluripotent Stem Cells
matrigel
Mus
Pericytes
Permeability
Plasma
Polystyrenes
Serum
Stem Cells
Sulfoxide, Dimethyl
Tissues
Versene
Animals
Biological Factors
Cells
Clone Cells
Collagen Type IV
Common Cold
Immunofluorescence
Intestines, Small
Liver
Lung
Lymphocyte
Mus
Novus
Rabbits
Spleen
Human enteroid cultures were established from biopsies obtained after endoscopic or surgical procedures utilizing the methods developed by the laboratory of Dr. Hans Clevers22 (link). De-identified biopsy tissue was obtained from healthy subjects provided informed consent at Johns Hopkins University and all methods were carried out in accordance with approved guidelines and regulations. All experimental protocols were approved by the Johns Hopkins University Institutional Review Board (IRB# NA_00038329). Briefly, enteroids generated from isolated intestinal crypts17 (link) were maintained as cysts embedded in Matrigel (Corning, USA) in 24-well plates and cultured in Wnt3A containing NDM. Once multiple enteroid cultures had been generated, multiple wells were pooled, triturated in Cultrex Organoid Harvesting Solution (Trevigen, USA), and the fragments were collected by centrifugation and resuspended in NDM. Enteroid fragments (100 μl) were added onto 0.4 μm or 1.0 μm pore transparent polyester (PET) membrane 24-well cell culture inserts (Transwell; Corning, USA or Millipore, USA) pre-coated with human collagen IV (30 μg/ml; Sigma-Aldrich, USA). NDM (600 μl) was added to the wells of the receiver plate, and the cultures incubated at 37 °C, 5% CO2. Under these conditions, enteroid cultures reached confluency in 7-14 days. Monolayer differentiation was induced by incubation in Wnt3A-free20 (link) and Rspo-1-free DFM for five days. Monolayer confluency and differentiation were monitored by measuring TER with an ohmmeter (EVOM2 (link); World Precision Instruments, USA). The unit area resistances (ohm*cm2) were calculated according to the growth surface area of the inserts; 0.33 cm2 for both 0.4 and 1.0 μm inserts. Enteroids were derived from segments of the small intestine (duodenum, n = 5; jejunum, n = 1) and proximal colon (n = 1) for Fig. 3c . For all other experiments, results were generated from duodenal and jejunal enteroids derived from intestinal biopsies of healthy subjects.
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Biopsy
Centrifugation
Collagen Type IV
Colon
Cyst
Duodenum
Endoscopy
Healthy Volunteers
Homo sapiens
Intestines
Intestines, Small
Jejunum
matrigel
Operative Surgical Procedures
Organoids
Plasma Membrane
Polyesters
Tissues
Protocol full text hidden due to copyright restrictions
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Cell Culture Techniques
Cells
Collagen Type IV
Endothelium
FN1 protein, human
Induced Pluripotent Stem Cells
matrigel
Phenotype
Y 27632
Adult
alpha 2-Plasmin Inhibitor
Antibodies, Blocking
Aves
benzamide
Bicarbonate, Sodium
Brain
Caspase 3
Cell Nucleus
Cells
Collagen Type IV
FASLG protein, human
Glucose
Gold
HEPES
Immunofluorescence
Innovativeness
Malignant Neoplasms
Mice, Nude
Mus
Penicillins
Plasminogen Activator Inhibitor 2
Sepharose
Sodium Chloride
Streptomycin
Sulfate, Magnesium
Tissue, Membrane
Tissues
Most recents protocols related to «Collagen Type IV»
hPIs in suspension and encapsulated in hydrogel maintained in culture for 1, 14 and 28 days were fixed in paraformaldehyde (PFA) 2% and 4% (w/v in PBS) and cryosectioned at 50 µm-thick via Cryostat (Histo-Line Laboratories). Sections were permeabilized with 0.3% Triton X-100 for 10 min at 4°C and blocked with 10% normal goat serum (NGS, Gibco) for 1 h at room temperature. The following primary antibodies were used: rabbit anti-insulin (1:300, ThermoFisher), mouse anti-chromogranin (1:100, ThermoFisher), mouse anti-glucagon (1:8000, Sigma-Aldrich), rabbit anti-Ki67 (1:750. Novus Biologicals), rabbit anti-vWF (1:500, DakoCytomation), mouse anti-fibroblast (1:200, Acris Antibodies), rabbit anti-collagen IV (1:100, Cedarlane), mouse anti-collagen I (1:2000, Sigma-Aldrich), rabbit anti-laminin (1:30, Sigma-Aldrich). To reveal primary antibodies, the following secondary antibodies were used: goat anti-rabbit Cy3 (1:1,000, Jackson), goat anti-mouse Cy3 (1:1,000, Jackson), goat anti-rabbit Alexa 488 (1:1,000, Invitrogen) and goat anti-mouse Alexa 488 (1:1,000, Invitrogen). Cell nuclei are stained with HOECHST 33342 (1:500, Molecular Probes).
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Antibodies
Biological Factors
Cell Nucleus
Chromogranins
Collagen Type I
Collagen Type IV
Fibroblasts
Glucagon
Goat
HOE 33342
Hydrogels
Insulin
Laminin
Molecular Probes
Mus
Novus
paraform
Rabbits
Serum
Triton X-100
Primary human bronchial epithelial cells (HBE) were provided by Nationwide Children’s Hospital Cure Cystic Fibrosis Columbus Epithelial Cell Core (Columbus, OH) without identifiers (exempt status from the Institutional Review Board). Primary HBE cells were cultured as previously reported in our lab [38 (link),70 (link)]. Briefly, primary human airway cells were seeded onto 6.5mm Transwell filters (Corning) coated with collagen Type IV (Sigma) with ~50,000 cells. Cells were seeded and fed with ROCK inhibitor-supplemented air-liquid interface (ALI) media on both sides of the membrane until optimal epithelial integrity was reached, as measured by an epithelial voltohmeter (~7 days; ~300 Ohms·cm2). Basal medium was replaced with PneumaCult-ALI (Stem Cell Technologies) differentiation medium, and cells were fed three times per week for ~3 weeks until fully differentiated. Cell differentiation and integrity of the bronchial epithelium were confirmed before each experiment. Cultures were also checked for absence of mycoplasma contamination.
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Bronchi
Bronchus, Primary
Cells
Collagen Type IV
Cystic Fibrosis
Differentiations, Cell
Epithelial Cells
Epithelium
Ethics Committees, Research
Homo sapiens
Mycoplasma
Stem Cells
Tissue, Membrane
The primary cultures of brain endothelial cells (BECs) were prepared from 6–7-month-old WT and APOB-100 transgenic mice as described in detail by Lénárt et al. [32 (link)]. 3 male and 3 female mice were used in both the WT (n = 6) and the APOB-100 group (n = 6) for each isolation. Forebrains were collected in ice-cold sterile PBS; meninges were removed, grey matter was minced by scalpel into 1 mm3 pieces and digested with 10 mg/ml collagenase II and 1 mg/ml DNase I in Dulbecco’s modified Eagle’s medium (DMEM)/F12 for 50 min at 37 °C. Microvessels were separated from myelin containing elements by centrifugation (1000×g, 20 min) in 20% bovine serum albumin (BSA)-DMEM and further digested with 10 mg/ml collagenase-dispase (Roche, Basel, Switzerland) and 1 mg/ml DNase I in DMEM/F12 for 35 min at 37 °C. Then they were washed twice in DMEM/F12 before plating on collagen type IV and fibronectin-coated (100 µg/ml each) dishes, 6 well plates (Corning Costar Co., Lowell, MA, USA) or cell culture inserts (Transwell clear, 1 cm2; pore size of 0.4 μm; Corning Costar Co.). Cultures were maintained in DMEM/F12 supplemented with 15% plasma-derived bovine serum (PDS; First Link, Wolverhampton, UK), 1 ng/ml basic fibroblast growth factor (Roche) and 100 μg/ml heparin. During the first 2 days, the culture medium contained puromycin (4 μg/ml) in order to selectively remove P-glycoprotein-negative contaminating cells [33 (link)]. Cultures reached confluency within a week and were used for experiments. To induce BBB characteristics, BECs were co-cultured with mouse astroglial cells. The resulting double co-culture model was used for permeability studies and transendothelial electrical resistance measurements [34 (link), 35 (link)].
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Apolipoprotein B-100
Astrocytes
Bos taurus
Brain
Cell Culture Techniques
Cells
Centrifugation
Coculture Techniques
Collagenase
Collagen Type IV
Common Cold
Culture Media
Deoxyribonuclease I
dispase
Eagle
Endothelial Cells
Endothelium
Females
Fibroblast Growth Factor 2
Fibronectins
Gray Matter
Heparin
Hyperostosis, Diffuse Idiopathic Skeletal
isolation
Males
Meninges
Mice, Transgenic
Microvessels
Mus
Myelin
P-Glycoprotein
Permeability
Plasma
Primary Cell Culture
Prosencephalon
Puromycin
Resistance, Electrical
Serum
Serum Albumin, Bovine
Sterility, Reproductive
Type II Mucolipidosis
In vitro human BBB models were developed by combining three immortalized cell lines [11 (link)]). Briefly, HBPC/ci37 cells were seeded on the bottom side of the collagen IV- and Fibronectin-coated polycarbonate membrane of a transwell insert (Millicell cell culture insert 24-well hanging inserts, 0.4 μm PET; Merck, Darmstadt, Germany) at a density of 1.0 × 104 cells/insert. The cells were then cultured for 1 day to allow them to attach firmly. HASTR/ci35 cells were seeded (5.0 × 104 cells/well) on collagen I-coated 24-well plates (Greiner Bio-one, Frickenhausen, Germany) and maintained in astrocyte culture medium. HBPC/ci37 cells were induced to differentiate by replacing the pericyte medium with pericyte differentiation medium, which was consisted of FBS- and blasticidin S-free pericyte medium; HASTR/ci35 cells were induced to differentiate by replacing the astrocyte culture medium with astrocyte differentiation medium, which was consisted of FBS- and blasticidin S-free astrocyte growth medium supplemented with 1 mM adenosine 3′,5′-cyclic monophosphate sodium salt monohydrate. After the differentiation media were added, both cell lines were cultured at 37 °C for 24 h. To start a coculture, HBMEC/ci18 cells were seeded on the inner side of the HBPC/ci37 cell culture insert at a density of 1.0 × 105 cells. Finally, the transwell inserts with HBMEC/ci18 cells and HBPC/ci37 cells were transferred into 24-well plates containing HASTR/ci35 cells. The cells were re-fed with VEGF- and EGF-free VascuLife complete medium in the inner insert and the Neurobasal medium with N2 supplement in the lower chamber. Day 0 was defined as the day of EC plating on the membrane. The cells were incubated at 33 °C.
On Day 1, the trans-endothelial electrical resistance (TEER) was measured by an EVOM2 voltohmmeter (World Precision Instruments, Sarasota, California, USA) with chopstick electrodes. The net resistance value was calculated by subtracting the measured resistance value of the insert membrane from the measured resistance value of the coculture. TEER (Ω × cm2) = the net resistance value (Ω) × surface area (cm2).
On Day 1, the trans-endothelial electrical resistance (TEER) was measured by an EVOM2 voltohmmeter (World Precision Instruments, Sarasota, California, USA) with chopstick electrodes. The net resistance value was calculated by subtracting the measured resistance value of the insert membrane from the measured resistance value of the coculture. TEER (Ω × cm2) = the net resistance value (Ω) × surface area (cm2).
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Adenosine
Astrocytes
blasticidin S
Cell Culture Techniques
Cell Lines
Cells
Coculture Techniques
Collagen Type I
Collagen Type IV
Dietary Supplements
Endothelium
Fibronectins
Homo sapiens
Pericytes
polycarbonate
Resistance, Electrical
Sodium
Sodium Chloride
Tissue, Membrane
Vascular Endothelial Growth Factors
HBMEC/ci18, HBVPC/ci37, and HASTR/ci35 were established and supplied by Prof. Furihata. VascuLife complete medium was purchased from Kurabo (Osaka, Japan). Astrocyte growth medium, Neurobasal medium, fibronectin, anti-TfR antibody (#13–6800), and rhodamine 123 were purchased from Thermo Fisher Scientific (Waltham, USA). Pericyte medium was purchased from ScienCell Research Laboratories (Carlsbad, CA, USA). Blasticidin S was purchased from Fujifilm Wako (Tokyo, Japan). Collagen IV and collagen I were purchased from Nitta Gelatin (Osaka, Japan). Anti-Claudin-5 (ab131259), anti-P-gp (ab170904), and anti-Glut1 (ab115730) antibodies were purchased from Abcam (Cambridge, UK). Anti-β-actin antibody was purchased from Sigma–Aldrich (A5316, St. Louis, MO, USA). Anti-CD31 antibody was purchased from Proteintech (66065-1-Ig, Rosemont, IL, USA). Anti-ZO-1 antibody was purchased from Invitrogen (#339100). Anti-BCRP antibody was purchased from Cell Signaling Technology (#4477, Danvers, MA, USA). Anti-rabbit IgG conjugated with Alexa Fluor 488 or 594, anti-goat IgG conjugated with Alexa Fluor 488, and anti-mouse IgG conjugated with Alexa Fluor 488 or 594 were purchased from Molecular Probes. Fetal bovine serum (FBS) and Dulbecco's modified Eagle's medium (DMEM) were purchased from Life Technologies (Grand Island, NY, USA). Can Get Signal was purchased from TOYOBO (Osaka, Japan). Hoechst 33,342 and DAPI were purchased from Dojindo (Tokyo, Japan). 2-NBDG was purchased from Cayman Chemical Company (Ann Arbor, Michigan, USA). Digoxin was purchased from Alfer Aeser (Heysham, Lancashire, UK). Dantrolene and salazosulfapyridine (sulfasalazine, SASP) were purchased from Tocris Bioscience (Minneapolis, MN, USA). Adenosine 3′,5′-cyclic monophosphate sodium salt monohydrate was purchased from Merck (Darmstadt, Germany). Both human transferrin with no conjugated fluorophore and human transferrin conjugated with Alexa Fluor 488 were purchased from Jackson ImmunoResearch (West Grove, USA).
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2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose
Actins
Adenosine
alexa fluor 488
anti-IgG
Antibodies
Antibodies, Anti-Idiotypic
Astrocytes
blasticidin S
Caimans
Claudin-5
Collagen Type I
Collagen Type IV
Dantrolene
DAPI
Digoxin
Fetal Bovine Serum
Fibronectins
Gelatins
Goat
Homo sapiens
Molecular Probes
Mus
Pericytes
Rabbits
Rhodamine 123
SLC2A1 protein, human
Sodium
Sodium Chloride
Sulfasalazine
Transferrin
TXN protein, human
Top products related to «Collagen Type IV»
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Collagen IV is a type of extracellular matrix protein found in the basement membrane of cells. It provides structural support and plays a role in cell adhesion and differentiation.
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Ab6586 is a mouse monoclonal antibody that detects the NOS2 protein. NOS2 is an enzyme involved in the production of nitric oxide. This antibody can be used in various immunoassay applications to detect and quantify NOS2 levels.
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Fibronectin is an extracellular matrix glycoprotein that plays a role in cell adhesion, growth, migration, and differentiation. It is a key component of the cellular microenvironment and is involved in various biological processes.
Sourced in United States, Switzerland, Belgium, Germany
Collagen type IV is a key structural component of basement membranes. It provides a platform for the assembly of other basement membrane proteins and plays a crucial role in the organization and function of this extracellular matrix.
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Laminin is a protein component found in the extracellular matrix of cells. It plays a key role in cell attachment, differentiation, and migration processes.
Sourced in United States, United Kingdom
Collagen IV is a major structural component of basement membranes, providing mechanical support and a scaffold for cell attachment. It is expressed in various tissues and plays a crucial role in cell adhesion, migration, and differentiation.
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Bovine serum albumin (BSA) is a common laboratory reagent derived from bovine blood plasma. It is a protein that serves as a stabilizer and blocking agent in various biochemical and immunological applications. BSA is widely used to maintain the activity and solubility of enzymes, proteins, and other biomolecules in experimental settings.
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Fetal Bovine Serum (FBS) is a cell culture supplement derived from the blood of bovine fetuses. FBS provides a source of proteins, growth factors, and other components that support the growth and maintenance of various cell types in in vitro cell culture applications.
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Fibronectin is a high molecular weight glycoprotein found in the extracellular matrix and plasma. It plays a crucial role in cell adhesion, growth, migration, and differentiation.
Sourced in United Kingdom, United States, China
Anti-collagen IV is a laboratory reagent used for the detection and quantification of collagen IV in biological samples. It is a highly specific antibody that binds to collagen IV, a key structural component of basement membranes. This product can be used in various immunoassay techniques to measure collagen IV levels.
More about "Collagen Type IV"
Collagen IV, a key structural component of basement membranes, plays a vital role in the integrity and functioning of various organ systems, including the kidneys, lungs, and blood vessels.
Researchers studying this important collagen type can leverage PubCompare.ai's AI-powered platform to optimize their research protocols and enhance reproducibility.
The platform can effortlessly locate the best protocols from literature, pre-prints, and patents, while providing AI-driven comparisons to identify the most effective products.
This can help take the guesswork out of research and elevate findings on Collagen Type IV.
Basement membranes are specialized extracellular matrices that provide support and organization to tissues throughout the body.
Collagen IV, a major constituent of these membranes, is essential for their structural integrity and function.
Researchers studying this collagen type can explore related terms such as Ab6586, Fibronectin, Laminin, Bovine serum albumin (BSA), and Fetal bovine serum (FBS) to enhance their understanding of the complex extracellular environment in which Collagen IV operates.
By leveraging PubCompare.ai's AI-powered platform, researchers can streamline their workflow, optimize their research protocols, and improve the reproducibility of their findings on Collagen Type IV.
The platform's ability to effortlessly locate the best protocols from various sources and provide AI-driven comparisons can help researchers make informed decisions and take the guesswork out of their work.
This, in turn, can elevate the quality and impact of their research on this important collagen type and its role in the body's organ systems.
Researchers studying this important collagen type can leverage PubCompare.ai's AI-powered platform to optimize their research protocols and enhance reproducibility.
The platform can effortlessly locate the best protocols from literature, pre-prints, and patents, while providing AI-driven comparisons to identify the most effective products.
This can help take the guesswork out of research and elevate findings on Collagen Type IV.
Basement membranes are specialized extracellular matrices that provide support and organization to tissues throughout the body.
Collagen IV, a major constituent of these membranes, is essential for their structural integrity and function.
Researchers studying this collagen type can explore related terms such as Ab6586, Fibronectin, Laminin, Bovine serum albumin (BSA), and Fetal bovine serum (FBS) to enhance their understanding of the complex extracellular environment in which Collagen IV operates.
By leveraging PubCompare.ai's AI-powered platform, researchers can streamline their workflow, optimize their research protocols, and improve the reproducibility of their findings on Collagen Type IV.
The platform's ability to effortlessly locate the best protocols from various sources and provide AI-driven comparisons can help researchers make informed decisions and take the guesswork out of their work.
This, in turn, can elevate the quality and impact of their research on this important collagen type and its role in the body's organ systems.