Immunofluorescence staining was performed to determine the surface area of the cell. H9C2 cells were infected with the indicated lentivirus for 24 hours, stimulated with phosphate‐buffered saline (PBS), angiotensin II (1 μmol/L) or iTAK1 NG25 (2.5 μmol/L, HY‐15434, MCE) for 48 hours under a condition of 37.0°C, 5% CO2 and fixed with 4% formaldehyde. After permeabilization with 0.1% Triton X‐100 in PBS and blocking with a 10% bovine serum albumin solution at room temperature, cells were immunostained with an α‐actinin antibody (1:100 dilution, 05‐384; Merck Millipore, Burlington, MA), followed by staining with a fluorescent secondary antibody (donkey anti‐mouse IgG [H+L] secondary antibody, 1:200, A21202; Invitrogen, Carlsbad, CA). Image‐Pro Plus 6.0 software was used to measure the surface area of the cell.
Donkey anti mouse igg h l secondary antibody
Manufactured by Thermo Fisher Scientific
Donkey anti-mouse IgG [H+L] secondary antibody is a laboratory reagent used for the detection and quantification of mouse immunoglobulins (IgG) in various immunoassays and other applications. It is produced by immunizing donkeys with mouse IgG and purifying the resulting antibodies.
Automatically generated - may contain errors
Lab products found in correlation
α actinin antibody, by Merck Group (2 mentions)
α actinin, by Merck Group (1 mentions)
G1101 500ml, by Wuhan Servicebio Technology (1 mentions)
Bah66 0100, by Equitech-Bio (1 mentions)
Ab63638, by Abcam (1 mentions)
A21202, by Thermo Fisher Scientific (1 mentions)
A21207, by Thermo Fisher Scientific (1 mentions)
Ab175429, by Abcam (1 mentions)
Donkey anti mouse igg h l, by Abcam (1 mentions)
7 protocols using donkey anti mouse igg h l secondary antibody
1
Measuring Cell Surface Area by Immunofluorescence
2
Cell Surface Area Analysis of NRCMs
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The cell surface area of the NRCMs was assessed via immunofluorescence staining. Briefly, the cardiomyocytes were treated with Ang II or PE for 48 h after infection with the corresponding adenoviruses for 24 h. The cells were subsequently fixed with 3.7% formaldehyde, permeabilized with 0.1% Triton X-100 in PBS for 5 min and stained with α-actinin (05-384, Merck Millipore, 1:100 dilution), followed by staining with a fluorescent secondary antibody (Donkey anti-Mouse IgG (H+L) Secondary Antibody, A21202, Invitrogen, 1:200). The surface areas were measured using Image-Pro Plus software, version 6.0.
3
Immunostaining of Cardiomyocytes for Actinin
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NRCMs were cultured for 24 h and fixed with 4% formaldehyde (G1101-500ML, Servicebio) for 30 min, then permeated with 0.2%Triton X-100 in PBS and blocked with 10% BSA (BAH66-0100, Equitech Bio) at 37.0 °C. The cells on slides were incubated with α-actinin antibody (05-384, Merck Millipore, 1:100 dilution), followed by staining with a fluorescent secondary antibody (donkey anti-mouse IgG [H + L] secondary antibody, A21202, Invitrogen, 1:200) and then the slides were mounted with an antifade mounting medium containing DAPI. Cell surface area was measured using image-Pro Plus 6.0 software.
4
Measuring NRCM Surface Area by Immunostaining
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Immunofluorescence staining was conducted to test the surface area of the NRCMs. The cells were infected with the indicated adenovirus for 24 hours, and then stimulated with PBS or Ang II (1 μmol/L) for 48 hours followed by fixed with 3.7% formaldehyde. Then, the cells were immunostained with an α‐actinin antibody (05‐384, Merck Millipore, 1:100 dilution), followed by staining with a fluorescent secondary antibody (donkey anti‐mouse IgG [H+L] secondary antibody, A21202, Invitrogen, 1:200) after permeabilization with 0.1% Triton X‐100 in PBS and blocking with a 10% BSA solution at room temperature. Image‐Pro Plus 6.0 software was used to measure the surface area of the cells.
5
Immunofluorescence staining of NRCMs
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Immunofluorescence staining was performed to determine cell surface area. NRCMs were infected with the indicated adenovirus for 24 h, stimulated with PBS or Ang II (1 μmol/L) for 48 h under conditions of 37.0 °C and 5% CO2, and fixed with 4% formaldehyde. After permeabilization with 0.1% Triton X-100 in PBS and blocking with a 10% bovine serum albumin solution at room temperature, the cells were immunostained with an α-actinin antibody (05-384, Merck Millipore, 1:100 dilution), followed by staining with a fluorescent secondary antibody (donkey anti-mouse IgG [H + L] secondary antibody, A21202 , Invitrogen, 1:200). Image-Pro Plus 6.0 software was used to measure cell surface area.
6
Cytoskeleton Analysis in Neonatal Rat Cardiomyocytes
7
Rex1 Immunofluorescence in ESCs
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A total of 1 × 106 cells (WT ESCs, passage number 29) were required for optimal performance. The cells were fixed by adding 100 µl of 4% PFA to each sample for 15 min and then permeabilized by adding 100 µl of PBS and 0.3% Triton-X 100. The cells were stained with primary and secondary antibodies. The primary antibody was anti-Rex1 antibody (Abcam, ab175429) and the secondary antibodies were donkey anti-mouse IgG (H + L) Secondary Antibody (Life Technologies, A16018) and donkey anti-mouse IgG H + L (TRITC) preadsorbed (Abcam, ab7058).
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