Paraffin-embedded rat lung tissue was sectioned, dewaxed, and rehydrated as described previously [22 (link)]. Hematoxylin and eosin (H&E) staining was performed according to the operation manual of a hematoxylin and eosin staining kit (Beyotime, CHN). IHC was performed as described [23 (link)]. After blocking nonspecific protein binding, the sections were treated with mouse anti-α-SMA (1:500, Boster, Wuhan, CHN), anti-p-PI3K/anti-PI3K (1:100, Abcam, USA), anti-p-Akt/anti-Akt (1:200, Abcam, UK), anti-p-mTOR/anti-beta-actin (1:100, Abcam, UK), anti-p-PKC/anti-PKC (1:250, Cell Signaling, USA), or anti-p-p44/42/anti-p44/42 antibodies (1:250, Cell Signaling, USA) followed by incubation for 2 h with goat anti-rabbit secondary antibodies (HPR, Boster, CHN); then, ABC horseradish peroxidase reagent (Boster, CHN) was added for 30 min at room temperature. The secondary antibody controls (negative control) were incubated in PBS instead of the primary antibody. Specimens were observed under a fluorescent inverted microscope (IX73-A22FL/PH; Olympus Corporation, Japan), and images were captured by using the Image-Pro Plus software (version 6.0). The average optical density (IOD/area) was used as a semiquantitative index for analysis of the positive signals.
Mouse anti α sma
Manufactured by Boster Bio
Mouse anti-α-SMA is a primary antibody used for the detection of alpha-smooth muscle actin (α-SMA) in immunohistochemistry and Western blot applications. α-SMA is a cytoskeletal protein expressed in various cell types, including vascular smooth muscle cells, myofibroblasts, and other contractile cells.
Automatically generated - may contain errors
Lab products found in correlation
Goat serum, by Solarbio (2 mentions)
Ix73 a22fl ph, by Olympus (1 mentions)
Hematoxylin and eosin staining kit, by Beyotime (1 mentions)
Image pro plus software, by Olympus (1 mentions)
Rabbit anti nrf2, by Santa Cruz Biotechnology (1 mentions)
Anti fluorescence quenching agent, by Abcam (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Triton x 100, by Avantor (1 mentions)
Paraformaldehyde (pfa), by Sinopharm (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Biosharp (1 mentions)
Alexa fluor 568, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 647 conjugated antibody, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (1 mentions)
Triton x 100, by Solarbio (1 mentions)
3 protocols using mouse anti α sma
1
Immunohistochemical Analysis of Rat Lung Tissue
2
Immunofluorescence Analysis of α-SMA and Nrf2 in Cells
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Cells were seeded onto glass slide beforehand. When the confluence reached 70%, the cells were fixed with 4% paraformaldehyde (Sinopharm) for 15 min and permeated with 0.1% Triton X‐100 (Amresco, Solon, OH) at room temperature for 30 min. After blocking with goat serum (Solarbio) for 15 min, the cells were incubated with mouse anti‐α‐SMA (1:400; Boster) and rabbit anti‐Nrf2 (1:200; Santa Cruz) at 4°C overnight. After rinsing with PBS, the cells were incubated with goat antirabbit immunoglobulin G (IgG) labeled with Cy3 (1:200, diluted with PBS; Invitrogen) at room temperature in the dark for 60 min. After being counterstained with 4′,6‐diamidino‐2‐phenylindole (DAPI) (Invitrogen, USA), the cells were mounted in the presence of anti‐fluorescence quenching agent (Abcam), observed and photographed with a fluorescence microscope (Olympus) at 400× magnification.
3
Immunofluorescence analysis of TEBVs
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Cells were fixed in 4% paraformaldehyde for 15 min, and the paraffin sections of TEBVs were dewaxed. After washing twice with PBS, 0.05% Triton X-100 (Solarbio, 30 min, room temperature) and 10% goat serum (Solarbio, 40 min, 37°C) in PBS were used for permeation and blocking, respectively. The samples were then incubated with antibodies at 4°C overnight. The primary antibodies were as follows: mouse anti-Gli (1:200, Novus Biologicals), mouse anti-MYH11 (1:200, Abcam), mouse anti–α-SMA (1:300, Boster), rabbit anti-thrombospondin (1:200, Abcam), rabbit anti-Runx2 (1:200, Bioss), and mouse anti-OPN (1:200, Bioss). The secondary antibodies were as follows: Alexa Fluor 488–, Alexa Fluor 568–, and Alexa Fluor 647–conjugated antibodies (Invitrogen). The nuclei were stained with DAPI (Biosharp) for 5 min at room temperature. Confocal microscopy images were taken and analyzed by the Olympus laser scanning confocal microscope (Olympus SpinSR, Japan) and cellSens software, respectively.
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