rabbit anti fast myosin heavy chain  (Servicebio Inc)

Journal: bioRxiv

Article Title: Mechanical constraints organize 3D tissues and orchestrate muscle differentiation

doi: 10.1101/2024.10.03.616457

Figure Lengend Snippet: a) Metabolic activity of C2C12 cells with or without nanoparticle (NP) 2 hours (D0) or one day (D1) after the incorporation ([Fe] = 2 mM for 30 min). Mean +/-SEM. b) Representative images of C2C12 after 7 days of differentiation with or without NP incorporation at day 0. Long myotubes are visible in the entire field of view in both conditions. c) Immunofluorescence representative images of C2C12 after 3 days of differentiation and incorporation of NP at day 0. Myh4 and nuclei are shown in green and blue respectively.

Article Snippet: After rinsing the membranes with TBS-Tween 20 buffer (TBST), the membranes were blocked for 1h in the EveryBlot blocking buffer (12010020, Biorad) and incubated overnight at 4°C with primary antibodies rabbit anti-Myf6 (sc-301, Santa Cruz Biotechnology), or mouse anti-fast myosin heavy chain (Myh4) (MF 20, DSHB) diluted at 1:1000 with 5% BSA.

Techniques: Activity Assay, Immunofluorescence

Journal: bioRxiv

Article Title: Mechanical constraints organize 3D tissues and orchestrate muscle differentiation

doi: 10.1101/2024.10.03.616457

Figure Lengend Snippet: a) Primer sequences used for RPLP0, Myod1, Myog, Myf6, Tnnt1, Tnnt3, Myh1, Myh3, Myh4 and Ckm. b) Expression levels are normalised by the mRNA levels at day 1 without NP incorporation (2D D1). RPLP0 was used as a housekeeping gene. Mean +/-SEM (N=4).

Article Snippet: After rinsing the membranes with TBS-Tween 20 buffer (TBST), the membranes were blocked for 1h in the EveryBlot blocking buffer (12010020, Biorad) and incubated overnight at 4°C with primary antibodies rabbit anti-Myf6 (sc-301, Santa Cruz Biotechnology), or mouse anti-fast myosin heavy chain (Myh4) (MF 20, DSHB) diluted at 1:1000 with 5% BSA.

Techniques: Expressing

Journal: bioRxiv

Article Title: Mechanical constraints organize 3D tissues and orchestrate muscle differentiation

doi: 10.1101/2024.10.03.616457

Figure Lengend Snippet: a) Myogenic marker expression levels, determined by quantitative reverse transcription polymerase chain reaction (RT-qPCR). Relative mRNA expression levels are measured for control spheroids at D1 and D3, and for stretched aggregates at D3. RPLP0 was used as a housekeeping gene. At least N = 6 for control spheroids with 4 independent experiments, N=4 to 8 in 4 to 8 independent experiments for stretched aggregates at D3 for each gene. Mean ± SEM. Myod1, myogenic differentiation 1, Myog, myogenin, Myf6, myogenic factor 6, Tnnt, troponin T, Ckm, creatin kinase M-type, Myh, myosin heavy chain; mRNA, messenger RNA; D, day. b) Western Blot of control spheroids at D1 and D3 and stretched aggregates at D3 for fast myosin heavy chain, Myh4 and myogenic factor 6, Myf6. α -tubulin is used as a loading control. c) Representative confocal images of a spheroid and stretched aggregates at D3. Immunostaining of troponin 1 protein (magenta) is superimposed with nuclei (cyan) in a transverse cryosection of a control spheroid (left), of a stretched aggregate (middle) and in a longitudinal cryosection of a stretched aggregate. Scalebar = 200 µm.

Article Snippet: After rinsing the membranes with TBS-Tween 20 buffer (TBST), the membranes were blocked for 1h in the EveryBlot blocking buffer (12010020, Biorad) and incubated overnight at 4°C with primary antibodies rabbit anti-Myf6 (sc-301, Santa Cruz Biotechnology), or mouse anti-fast myosin heavy chain (Myh4) (MF 20, DSHB) diluted at 1:1000 with 5% BSA.

Techniques: Marker, Expressing, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Control, Western Blot, Immunostaining