ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)
Overview
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Product name
ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes -
Parental Cell Line
iPSC -
Organism
Human -
Tested applications
Suitable for: ICC, WB, RT-PCR, High throughput screening, Functional Studiesmore details -
Biosafety level
1 -
General notes
Introducing the ioSkeletal Myocytes, generated from human induced pluripotent stem cells (iPSCs) using opti-ox, a precise cellular reprogramming technology. Human stem cells, within days, convert into consistent and reliable skeletal myocytes, providing a high-quality human model for research, disease modelling and HTS.
ioSkeletal Myocytes demonstrate robust expression of key proteins of myofilaments (Desmin, Titin, Troponin, Myosin Heavy Chain, Dystrophin), coupled with the transition from immature (MYH3 and MYH8) to mature myosin heavy chain isoforms (MYH1) in a time dependent manner. By Day 10 post revival, skeletal myocytes form striated multinucleated myocytes that contract in response to acetylcholine.
Human skeletal myocytes are available at scale, easy to culture and ready for experiments within days, providing a reliable model for the study of muscle, neuromuscular and associated metabolic disorders.
In partnership with bit.bio
Karyotype: Normal
Seeding Density: 100,000 cells/cm2
Seeding compatibility: 6-, 12-, 24- and 96-well compatible
Quality control: Sterility, ICC and gene expression analysis
Research applications: Muscle research, Neuromuscular junction Research, Metabolic research, Drug development, Genetic screening (e.g. CRISPR screening), Contractions assays
This product is subject to limited use licenses from iPS Academia Japan Inc, TET Systems GmbH, ERS Genomics Limited and Sigma-Aldrich Co. LLC and is developed with Bit Bio patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
Properties
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Number of cells
Small - 2.5x106 cells/vial; Large - 5x106 cells/vial -
Viability
>85% -
Cell type
skeletal myocyte -
Gender
Male -
Mycoplasma free
Yes -
Storage instructions
Shipped on Dry Ice. Store in liquid nitrogen. -
Storage buffer
Constituent: 10% DMSO -
Research areas
Associated products
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Corresponding Antibody
- Anti-Cardiac Troponin T antibody [1F11] (ab10214)
- Anti-Myogenin antibody [EPR4789] (ab124800)
- Anti-Desmin antibody - Cytoskeleton Marker (ab15200)
- Anti-alpha Actinin/ACTN1 antibody [AT6/172] (ab18061)
- Anti-Myogenin antibody [F5D] (ab1835)
- Anti-Cardiac Troponin T antibody [EPR20266] (ab209813)
- Anti-Desmin antibody [Y66] - Cytoskeleton Marker (ab32362)
- Anti-Fast Myosin Skeletal Heavy chain antibody [MY-32] (ab51263)
- Anti-alpha Actinin/ACTN1 antibody [EP2527Y] (ab68194)
- Anti-Cardiac Troponin T antibody [1C11] (ab8295)
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab277612 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| ICC |
Use at an assay dependent concentration.
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| WB |
Use at an assay dependent concentration.
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| RT-PCR |
Use at an assay dependent concentration.
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| High throughput screening |
Use at an assay dependent concentration.
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| Functional Studies |
Use at an assay dependent concentration.
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| Notes |
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ICC
Use at an assay dependent concentration. |
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WB
Use at an assay dependent concentration. |
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RT-PCR
Use at an assay dependent concentration. |
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High throughput screening
Use at an assay dependent concentration. |
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Functional Studies
Use at an assay dependent concentration. |
Images
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)
High purity skeletal myocytes express myofilament proteins.
Immunofluorescence staining at day 10 post revival demonstrates robust expression of components of the contractile apparatus such as Desmin (top left), Dystrophin (top right), and Myosin Heavy Chain (bottom left), along with the muscle transcription factor Myogenin (bottom left). Cells also demonstrate expression of Troponin with visible striated fibres and multinucleation (bottom right).
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Functional Studies - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Cells demonstrate timewise gene expression of key myogenic markers.
ioSkeletal Myocytes gene expression. Following reprogramming, ioSkeletal Myocytes downregulate expression of pluripotency markers (A), and begin to express myosin heavy chain isoforms MYH3 and MYH8 (B). Through continued culture, ioSkeletal Myocytes demonstrate expression of mature myosin isoforms MYH7 and MYH1, along with DESMIN, DYSTROPHIN, MYOGENIN, and TITIN (C). Gene expression levels assessed by RT-qPCR (data expressed relative to parental iPSC, normalised to PBGD). Data represents day (Dx) post-thaw. -
Cell Culture - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)ioSkeletal Myocytes are available in two vial sizes, tailored to suit your experimental needs with minimal waste.
- Recommended seeding density for ioSkeletal Myocytes is 100,000 cells/cm2.
- One Small vial (2.5 x 106 viable cells) can plate a minimum of 0.5 x 24-well plate, 0.75 x 96-well plate, or 1 x 384-well plate.
- One Large vial (5 x 106 viable cells) can plate a minimum of 1 x 24-well plate, 1.5 x 96-well plate, or 2 x 384-well plates. -
Cell Culture - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Cells demonstrate classical myocyte morphology.
Day 1 to 10 post-thawing; 4X magnification; scale bar: 800µm
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Fast Myosin Skeletal Heavy chain using ab51263 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 (left panel), 5 (middle panel) and 10 days (right panel) post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab51263 at 5 µg/mL and ab6046, rabbit polyclonal to beta Tubulin, at 1/1000 dilution. Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150088, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab51263 gave comparable results using 4% formaldehyde fixation (10 min).
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Cardiac Troponin T using ab8295 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab8295 at 5 µg/mL and ab6046, rabbit polyclonal to beta Tubulin, at 1/1000 dilution. Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150088, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab8295 gave comparable results using MeOH fixation (100%, 5 min).
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Myogenin using ab124800 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 days post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab124800 at 0.5 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab124800 gave comparable results using MeOH fixation (100%, 5 min).
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Myogenin using ab1835 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 days post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab1835 at 1 µg/mL and ab6046, rabbit polyclonal to beta Tubulin, at 1/1000 dilution. Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150088, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab1835 gave comparable results using MeOH fixation (100%, 5 min).
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Actinin/ACTN1 using ab68194 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 (left panel), 5 (middle panel) and 10 days (right panel) post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab68194 at 0.5 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Actinin/ACTN1 using ab68194 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 (left panel), 5 (middle panel) and 10 days (right panel) post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab68194 at 0.5 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Actinin/ACTN1 using ab18061 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab18061 at 5 µg/mL and ab6046, rabbit polyclonal to beta Tubulin, at 1/1000 dilution. Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150088, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Cardiac Troponin T using ab10214 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab10214 at 1 µg/mL and ab6046, rabbit polyclonal to beta Tubulin, at 1/1000 dilution. Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150088, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab10214 also gave a positive staining using 4% formaldehyde fixation (10 min).
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Cardiac Troponin T using ab209813 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab209813 at 0.02 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab209813 also gave a positive staining using MeOH fixation (100%, 5 min).
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Desmin using ab32362 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab32362 at 0.02 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab32362 gave comparable results using 4% formaldehyde fixation (10 min).
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of Desmin using ab15200 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 10 days post induction.
The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab15200 at 0.1 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
The antibody ab15200 gave comparable results using MeOH fixation (100%, 5 min).
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Immunocytochemistry - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Critically for metabolic studies, data demonstrates expression of the insulin regulated glucose transporter GLUT4 (Part 2)
Immunocytochemistry at Day 7 post-revival demonstrates expression of GLUT4 in peri-nuclear regions, and striations, in the ioSkeletal Myocytes.
Image courtesy of Dougall Norris & Daniel Fazakerley, Wellcome-MRC Institute of Metabolic Science.
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RT-PCR - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Critically for metabolic studies, data demonstrates expression of the insulin regulated glucose transporter GLUT4 (Part 1)
RT-qPCR at Day 10 post-revival demonstrates expression of GLUT4 in the ioSkeletal Myocytes, compared to undifferentiated hiPSCs and ioGlutamatergic Neurons.
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Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Immunofluorescence staining of GLUT4 using ab33780 in ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612), which were differentiated for 3 (left panel), 5 (middle panel) and 10 days (right panel) post induction.
The cells were fixed with 100% MeOH (5 min) and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab33780 at 5 µg/mL and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown. Gamma is adjusted to 1.5 in all channels.
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Western blot - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)Critically for metabolic studies, data demonstrates expression of the insulin regulated glucose transporter GLUT4 (Part 3)
Western blotting of differentiated 3T3-L1 adipocytes and maturing ioSkeletal Myocytes demonstrates GLUT4 expression in a time-dependent manner.
Image courtesy of Dougall Norris & Daniel Fazakerley, Wellcome-MRC Institute of Metabolic Science.
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Cell Culture - ioSkeletal Myocytes - Human iPSC-Derived Skeletal Myocytes (ab277612)ioSkeletal Myocytes cells arrive ready to plate. A simple one-medium protocol generates fully differentiated and mature skeletal myocytes.
The 3 phase protocol for generating ioSkeletal Myocytes:
1. Induction (carried out at bit.bio)
2. Stabilization for 3 days with Doxycycline
3. Maintenance during which the skeletal myocytes mature.
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab277612 has not yet been referenced specifically in any publications.