ioSkeletal Myocytes - Human iPSC derived cells
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- ICC
Lab
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Actinin/ACTN1 using ab18061 in 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.
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Cardiac Troponin T using ab209813 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).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Actinin/ACTN1 using ab68194 in 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.
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Desmin using ab15200 in 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).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Myogenin using ab124800 in 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).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (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).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Actinin/ACTN1 using ab68194 in 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.
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Desmin using ab32362 in 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).
- ICC
Lab
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Fast Myosin Skeletal Heavy chain using ab51263 in 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).
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of GLUT4 using ab33780 in 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.
- ICC
Lab
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Cardiac Troponin T using ab8295 inab277612, 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).
- FuncS
Supplier Data
Functional Studies - ioSkeletal Myocytes - Human iPSC derived cells (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.
- ICC
Supplier Data
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (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).
- ICC
Lab
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Immunofluorescence staining of Cardiac Troponin T using ab10214 in 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).
- ICC
Supplier Data
Immunocytochemistry - ioSkeletal Myocytes - Human iPSC derived cells (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.
- WB
Supplier Data
Western blot - ioSkeletal Myocytes - Human iPSC derived cells (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.
false
- RT-PCR
Supplier Data
RT-PCR - ioSkeletal Myocytes - Human iPSC derived cells (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.
- Cell Culture
Supplier Data
Cell Culture - ioSkeletal Myocytes - Human iPSC derived cells (AB277612)
Cells demonstrate classical myocyte morphology.
Day 1 to 10 post-thawing; 4X magnification; scale bar : 800μm
- Cell Culture
Supplier Data
Cell Culture - ioSkeletal Myocytes - Human iPSC derived cells (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
Supplier Data
Cell Culture - ioSkeletal Myocytes - Human iPSC derived cells (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.
Reactivity data
Product details
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 and storage information
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Handling procedures
Culture medium
Refer to protocol booklet
Cell culture
Biosafety level
EU: 1 US: 1
Gender
Male
Viability
> 85%
Product protocols
- Visit the General protocols
- Visit the Troubleshooting
- Download websiteProtocolBooklet|en
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