The procedure to induce early stages of retinal differentiation was based on a previously described protocol with major modifications14 (link),22 (link),29 (link). Briefly, on day 0 (D0) of differentiation, hiPSC were enzymatically detached by dispase treatment, dissociated into small clumps, and cultured in suspension with mTeSR1 medium and 10 µM Blebbistatin (Sigma) to induce aggregate formation. Aggregates were gradually transitioned into neural-induction medium (NIM) containing Dulbecco’s modified eagle medium (DMEM)/F12 (1:1), 1% N2 supplement (Invitrogen), 1x minimum essential media-non essential amino acids (NEAA), 2µg ml−1 heparin (Sigma), by replacing the medium with a 3:1 ratio of mTeSR1/NIM on D1, 1:1 on D2, and 100% NIM on D3. On D7, aggregates (average size of 0.22 ± 0.05 mm) were seeded onto Matrigel (growth-factor-reduced; BD Biosciences) coated dishes containing NIM at an approximate density of 20 aggregates per cm2 (link), and switched to DMEM/F12 (3:1) supplemented with 2% B27 (without vitamin A, Invitrogen), 1x NEAA, and 1% antibiotic-antimycotic (Gibco) on D16. Thereafter, the medium was changed daily.
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Blebbistatin
Blebbistatin
Blebbistatin is a small-molecule inhibitor of non-muscle myosin II, a key regulator of cell motility and contractility.
It has been widely used in biomedical research to study the role of myosin II in diverse cellular processes, such as cell migration, cytokinesis, and organelle transport.
Blebbistatin selectively binds to the myosin II motor domain, locking it in a low-affinity state and preventing ATP hydrolysis, thereby inhibiting myosin II activity.
This makes it a valuable tool for investigating the function of myosin II in living cells and tissues.
Optimzie your Blebbistatin research with PubCompare.ai's AI-driven platform, which helps you easily locate protocols from literature, pre-prints, and patents, and use powerful comparison tools to identify the best protocols and products.
Improve your research reproducibility and efficiency today.
It has been widely used in biomedical research to study the role of myosin II in diverse cellular processes, such as cell migration, cytokinesis, and organelle transport.
Blebbistatin selectively binds to the myosin II motor domain, locking it in a low-affinity state and preventing ATP hydrolysis, thereby inhibiting myosin II activity.
This makes it a valuable tool for investigating the function of myosin II in living cells and tissues.
Optimzie your Blebbistatin research with PubCompare.ai's AI-driven platform, which helps you easily locate protocols from literature, pre-prints, and patents, and use powerful comparison tools to identify the best protocols and products.
Improve your research reproducibility and efficiency today.
Most cited protocols related to «Blebbistatin»
Amino Acids, Essential
Antibiotics
blebbistatin
dispase
Eagle
Growth Factor
Heparin
Human Induced Pluripotent Stem Cells
Hyperostosis, Diffuse Idiopathic Skeletal
matrigel
Nervousness
Retina
Vitamin A
Adenovirus Vaccine
blebbistatin
Cells
Flavoproteins
Fluorescence
Heart
Heart Ventricle
Langendorff Perfused Heart
Mice, Transgenic
Microfilaments
Microscopy, Confocal
Mitochondria
Muscle Cells
Myocardium
NADH
Neurons
Open Reading Frames
Pulse Rate
Reading Frames
Retention (Psychology)
rhod-2
Submersion
Superoxides
1-(3-sulfonatopropyl)-4-(beta)(2-(di-n-butylamino)-6-naphthylvinyl)pyridinium betaine
Actins
Action Potentials
Adenosine Triphosphatases
blebbistatin
Cerebral Ventricles
Chronic multifocal osteomyelitis
CM 2-3
Cold Temperature
Diastole
Dietary Supplements
Electricity
Endocardium
Fluorescent Dyes
Heart
Heart Arrest, Induced
Heart Atrium
Heart Ventricle
Homo sapiens
Light
Mammals
Membrane Potentials
Microelectrodes
Molecular Probes
Myocardial Contraction
Myosin Type II
Protein Isoforms
Pulse Rate
Reading Frames
Tissues
Vision
The procedure to induce early stages of retinal differentiation was based on a previously described protocol with major modifications14 (link),22 (link),29 (link). Briefly, on day 0 (D0) of differentiation, hiPSC were enzymatically detached by dispase treatment, dissociated into small clumps, and cultured in suspension with mTeSR1 medium and 10 µM Blebbistatin (Sigma) to induce aggregate formation. Aggregates were gradually transitioned into neural-induction medium (NIM) containing Dulbecco’s modified eagle medium (DMEM)/F12 (1:1), 1% N2 supplement (Invitrogen), 1x minimum essential media-non essential amino acids (NEAA), 2µg ml−1 heparin (Sigma), by replacing the medium with a 3:1 ratio of mTeSR1/NIM on D1, 1:1 on D2, and 100% NIM on D3. On D7, aggregates (average size of 0.22 ± 0.05 mm) were seeded onto Matrigel (growth-factor-reduced; BD Biosciences) coated dishes containing NIM at an approximate density of 20 aggregates per cm2 (link), and switched to DMEM/F12 (3:1) supplemented with 2% B27 (without vitamin A, Invitrogen), 1x NEAA, and 1% antibiotic-antimycotic (Gibco) on D16. Thereafter, the medium was changed daily.
Amino Acids, Essential
Antibiotics
blebbistatin
dispase
Eagle
Growth Factor
Heparin
Human Induced Pluripotent Stem Cells
Hyperostosis, Diffuse Idiopathic Skeletal
matrigel
Nervousness
Retina
Vitamin A
hESCs were dissociated to single cells and plated on Matrigel or Synthemax II‐SC Substrate (Corning, USA, https://www.corning.com ) coated plates at a density of 52.6 K/cm2 in mTeSR1 with 5 µM blebbistatin, a time point designated as day minus 1 (d‐1). Unless otherwise specified, a Matrigel cover layer was not added to the cultures after plating. One day after plating, mTeSR1 was completely exchanged for N2B27 media [1:1 mix of DMEM/F12 and Neurobasal with 1× GlutaMAX Supplement, 1× antibiotic‐antimycotic, 1% N2 Supplement, and 2% B27 Supplement (all from ThermoFisher Scientific)] to start differentiation; this day was designated as day 0 (d0). Small molecules were added to the cells on day 1 (d1), 24 hours after d0. Small molecule addition was done in fresh N2B27 media. Cells were fed with a full exchange of N2B27 media every other day unless a small molecule was to be removed or added on that day of differentiation, requiring daily feeding. The following small molecules were aliquoted as 1,000× stocks in dimethyl sulfoxide (DMSO) and used at the working concentration noted in parentheses: Forskolin (FSK; 25 µM—Cell Signaling Technology, Danvers, MA, https://www.cellsignal.com ), Dorsomorphin (1 µM—R&D Systems, Minneapolis, MN, https://www.rndsystems.com ), IDE2 (2.5 µM—R&D Systems), DAPT (10 µM—Cell Signaling Technology), LDN‐193189 (0.5 µM—Stemgent, Lexington, MA, https://www.stemgent.com ), SB431542 (10 µM—Sigma‐Aldrich). Nicotinamide (NIC; Sigma‐Aldrich) was resuspended in water at 100× and used at a 10 mM working concentration. Noggin (ThermoFisher Scientific) was resuspended in 10 mM acetic acid with 0.5% bovine serum albumin (BSA) for a 1,000× stock and used at 100 ng/ml. All small molecules were added as indicated. Specifically, for our DIDNF+D protocol, Dorsomorphin and IDE2 (DID) were added from day 1 to 6, NIC from day 1 to 10, (FSK from day 1 to 30, and DAPT from day 18 to 30. Differentiation was carried out at 37°C in 5%CO2/20% O2.
1,2-dilinolenoyl-3-(4-aminobutyryl)propane-1,2,3-triol
4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide
Acetic Acid
Antibiotics
blebbistatin
Cells
Colforsin
Dietary Supplements
dorsomorphin
Human Embryonic Stem Cells
LDN 193189
matrigel
Niacinamide
noggin protein
Serum Albumin, Bovine
Sulfoxide, Dimethyl
Most recents protocols related to «Blebbistatin»
Myosin II inhibitor (-) Blebbistatin (Abcam) was dissolved in dimethyl sulfoxide (DMSO) at a stock concentration of 5 mM. Spheroids were incubated in a dilution concentration of 10 µM for at least 3 hours before experiments.
1 mM of Lat-A stock (L12370, Thermo Fisher Scientific) was prepared by reconstituting the chemical in DMSO. The embryos were treated with Lat-A at a final concentration of 2 nM. 1 mM of Blebbistatin stock (13013–5, Enco) was prepared by reconstituting the chemical in DMSO. The embryos were treated with Blebbistatin at a final concentration of 2 µM. For the double inhibition experiments, embryos were treated with Blebb at a final concentration of 1.5 µM, and with Lat-A at a final concentration of 2 nM.
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To change the contractility of the cytoskeleton, the treatment of the samples with three different drugs was performed starting at day 4 after seeding until the end of the experiment: myosin II inhibitor Blebbistatin to inhibit the myosin–actin contractility; TGF-β1 to enhance the contractility and stabilize the cytoskeleton or Latrunculin A to inhibit actin assembly. (−)-Blebbistatin (Sigma-Aldrich Chemie GmbH) was added to the growth media at a final concentration of 2 µM with 0.1% DMSO (Sigma-Aldrich Chemie GmbH). The recombinant human TGF-β1 (Invitrogen, MD, USA) was added at a final concentration of 1 ng/mL. The actin monomer-binding toxin Latrunculin A (Millipore, Darmstadt, Germany) was added at a final concentration of 20 nM with 0.1% DMSO. To exclude the effect of the solvent DMSO, additional samples were treated with 0.1% DMSO in the same period.
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Cells were plated and grown as previously described to produce primarily Intermediate architectures [8 (link)]. After 24 hrs, media was removed and cells were washed with PBS, after which cells were incubated for the specified time in magnesium-calcium-free PBS supplemented with 2 mM Magnesium dichloride. We found that for treatments of up to 3 hours in calcium free media, cell monolayers remained intact and mostly confluent, though there were occasionally cell scale gaps that would develop in the monolayer. These small gaps are unlikely to affect classification by ALAn and are unlikely to impact average architecture of the tissue.
For blebbistatin treated monolayers, cells were grown to an Intermediate architecture and treated with 50 μg/ml of blebbistatin or the equivalent amount of DMSO as a control for the specified time. After treatments cells were fixed and stained as per the above protocol.
For blebbistatin treated monolayers, cells were grown to an Intermediate architecture and treated with 50 μg/ml of blebbistatin or the equivalent amount of DMSO as a control for the specified time. After treatments cells were fixed and stained as per the above protocol.
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(-)-Blebbistatin (Selleckchem, S7099) was dissolved in DMSO to 10 mM and applied to cells in DMEM at 50 µM for 1 h. For wash-out, the DMEM was replaced with fresh serum-containing culture medium for 15 min.
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Top products related to «Blebbistatin»
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Blebbistatin is a small molecule that selectively inhibits non-muscle myosin II ATPase activity. It is commonly used as a research tool in cell biology and biochemistry studies.
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Nocodazole is a synthetic compound that acts as a microtubule-destabilizing agent. It functions by binding to and disrupting the polymerization of microtubules, which are essential components of the cytoskeleton in eukaryotic cells. This property makes Nocodazole a valuable tool in cell biology research for studying cell division, cell motility, and other cellular processes that rely on the dynamics of the microtubule network.
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Blebbistatin is a small-molecule inhibitor that selectively blocks the ATPase activity of non-muscle myosin II. It is commonly used in cell biology and biochemistry research to study the role of myosin II in various cellular processes.
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DMSO is a versatile organic solvent commonly used in laboratory settings. It has a high boiling point, low viscosity, and the ability to dissolve a wide range of polar and non-polar compounds. DMSO's core function is as a solvent, allowing for the effective dissolution and handling of various chemical substances during research and experimentation.
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Blebbistatin is a small molecule that selectively inhibits non-muscle myosin II. It is a useful tool for investigating the role of myosin II in various cellular processes.
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Latrunculin A is a chemical compound used in laboratory research. It functions as a potent inhibitor of actin polymerization, disrupting the cytoskeleton in cells. Latrunculin A is commonly utilized in cell biology studies to investigate the role of the actin cytoskeleton in various cellular processes.
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CK-666 is a laboratory equipment product manufactured by Merck Group. It is a small molecule inhibitor that disrupts the function of the Arp2/3 complex, which is involved in the regulation of actin cytoskeleton dynamics. The core function of CK-666 is to serve as a research tool for studying cellular processes related to the Arp2/3 complex.
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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.
More about "Blebbistatin"
Blebbistatin is a small-molecule inhibitor of non-muscle myosin II, a key regulator of cellular processes like motility, contractility, migration, cytokinesis, and organelle transport.
It works by selectively binding to the myosin II motor domain, locking it in a low-affinity state and preventing ATP hydrolysis, thereby inhibiting myosin II activity.
This makes blebbistatin a valuable research tool for investigating the function of myosin II in living cells and tissues.
To optimize your blebbistatin research, consider utilizing PubCompare.ai's AI-driven platform.
This platform can help you easily locate relevant protocols from literature, preprints, and patents, and provides powerful comparison tools to identify the best protocols and products.
This can improve your research reproducibility and efficiency.
Other small-molecule inhibitors like Y-27632 (a ROCK inhibitor), cytochalasin D (an actin polymerization inhibitor), nocodazole (a microtubule depolymerizer), and latrunculin A (another actin polymerization inhibitor) can also be useful for studying the cytoskeleton and related cellular processes.
Similarly, the small-molecule CK-666 inhibits the Arp2/3 complex, which is involved in actin nucleation and branching.
When working with these compounds, it's important to consider the vehicle (often DMSO) and serum conditions (e.g., FBS) that may affect their activity and cellular responses.
Careful experimental design and troubleshooting can help ensure the reliability and reproducibility of your research findings.
It works by selectively binding to the myosin II motor domain, locking it in a low-affinity state and preventing ATP hydrolysis, thereby inhibiting myosin II activity.
This makes blebbistatin a valuable research tool for investigating the function of myosin II in living cells and tissues.
To optimize your blebbistatin research, consider utilizing PubCompare.ai's AI-driven platform.
This platform can help you easily locate relevant protocols from literature, preprints, and patents, and provides powerful comparison tools to identify the best protocols and products.
This can improve your research reproducibility and efficiency.
Other small-molecule inhibitors like Y-27632 (a ROCK inhibitor), cytochalasin D (an actin polymerization inhibitor), nocodazole (a microtubule depolymerizer), and latrunculin A (another actin polymerization inhibitor) can also be useful for studying the cytoskeleton and related cellular processes.
Similarly, the small-molecule CK-666 inhibits the Arp2/3 complex, which is involved in actin nucleation and branching.
When working with these compounds, it's important to consider the vehicle (often DMSO) and serum conditions (e.g., FBS) that may affect their activity and cellular responses.
Careful experimental design and troubleshooting can help ensure the reliability and reproducibility of your research findings.