Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human skeletal muscle tissue labelling Desmin with purified ab32362 at 1/2000. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

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).

All lanes : Anti-Desmin antibody [Y66] - Cytoskeleton Marker (ab32362) at 1/500000 dilution (purified)

Lane 1 : Human skeletal muscle tissue lysate
Lane 2 : Human fetal heart tissue lysate
Lane 3 : Human fetal muscle tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 53 kDa
Observed band size: 53 kDa

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

Unpurified ab32362 staining Desmin (green) in Human skeletal muscle cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with methacarn and blocked with 10% serum for 20 minutes at 22°C. Samples were incubated with primary antibody (1/150) for 12 hours. An Alexa Fluor^® 488-conjugated Goat anti-rabbit IgG polyclonal (1/200) was used as the secondary antibody. Blue - DAPI-nuclei. Red - WGA. 40X objective.

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Immunocytochemistry/Immunofluorescence analysis of C2C12 (Mouse myoblasts myoblast) cells labeling Desmin with ab32362 at 1/500. Cells were fixed with 100% Methanol. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. 

 

ab195889, Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 was used as counterstain antibody.

Confocal image showing cytoplasmic staining on C2C12 cell line.

 

Flow Cytometry analysis of C2C12 cells labelling Desmin with purified ab32362 at 1/70 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

Immunocytochemistry/Immunofluorescence analysis of A673 cells labelling Desmin with purified ab32362 at 1/50. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.

Control: primary antibody (1/50) and secondary antibody, ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500).

Immunofluorescent analysis of Human mitochondria injected rabbit hearts sections stained for Desmin (Green) using ab32362. MTCO2, the human-specific mitochondrial marker was stained in red, and the nuclei was stained using the DNA stain DAPI (blue).

All lanes : Anti-Desmin antibody [Y66] - Cytoskeleton Marker (ab32362) at 1/500000 dilution (purified)

Lane 1 : Mouse heart tissue lysate
Lane 2 : Rat heart tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 53 kDa
Observed band size: 53 kDa

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

All lanes : Anti-Desmin antibody [Y66] - Cytoskeleton Marker (ab32362) at 1/100000 dilution

Lane 1 : Guinea pig heart tissue lysate
Lane 2 : Guinea pig muscle tissue lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

Predicted band size: 53 kDa
Observed band size: 53 kDa

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

All lanes : Anti-Desmin antibody [Y66] - Cytoskeleton Marker (ab32362) at 1/500 dilution (unpurified)

All lanes : Human skeletal muscle whole tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : IRDye® 680-conjugated anti-rabbit at 1/5000 dilution

Developed using the ECL technique.

Performed under reducing conditions.

Predicted band size: 53 kDa
Observed band size: 53 kDa


Exposure time: 50 seconds

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Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of normal human uterus tissue labelling Desmin with unpurified ab32362.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of normal human urinary bladder tissue labelling Desmin with unpurified ab32362.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of human skeletal muscle tissue labelling Desmin with unpurified ab32362.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of normal human brain tissue. Unpurified ab32362 shows negative staining.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of normal human tonsil tissue. Unpurified ab32362 shows negative staining.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Unpurified ab32362 staining Desmin in nude rat esophagheal tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer pH 6. Samples were then blocked with 1% BSA for 20 minutes at 25°C and then incubated with unpurified ab32362 at a 1/400 dilution for 16 hours at 25°C. The secondary used was an undiluted goat anti-rabbit HRP conjugated polyclonal.Striated muscle cells of muscular propria are strongly positive for desmin. Smooth muscle cells and vascular smooth muscle cells in submucosal layer are also positive for desmin.

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Unpurified ab32362 staining Desmin (green) in Mouse aorta smooth muscle cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formalin and blocked with 10% serum for 20 minutes at 22°C. Samples were incubated with primary antibody (1/150) for 1 hour at 22°C. An Alexa Fluor^® 488-conjugated Goat anti-rabbit IgG polyclonal (1/200) was used as the secondary antibody. Blue - nuclei.

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