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2 protocols using antigoat igg hrp conjugated antibody

1

Western Blot Analysis of Cellular Proteins

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Cells were lysed, and samples (40 µg protein per lane) were subjected to electrophoresis on 7 % (MPR), 10 % (clathrin), or 12 % (LC3) SDS-polyacrylamide gel followed by transfer to a polyvinylidene difluoride membrane. Membranes were blocked overnight with 5 % BSA and then incubated with appropriate primary antibodies overnight at 4 °C followed by incubation with antigoat IgG HRP-conjugated antibody (Santa Cruz) for 1 h at room temperature. The bands were visualized by ECL Western plus kit (GE healthcare). To confirm equal loading of protein, the membrane were also probed for cadherin, tubulin, or β-actin.
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2

Vascular Smooth Muscle Cell Characterization

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Collagenase type II, Elastase, and Exisulind were obtained from Sigma-Aldrich co. (St Louis, MO, USA). PDGF-BB was purchased from R&D systems (Minneapolis, MN, USA). Goat anti-PKG I, rabbit anti-VE-cadherin, mouse anti-osteoponin, rabbit anti-CD31, and goat anti-actin antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Rabbit anti-VASP (Ser239), rabbit anti-t-Akt, and mouse anti-p-Akt (Ser473) were purchased from Cell Signaling (Berkely, MA, USA). Mouse anti-calponin antibody was obtained from Sigma-Aldrich co. (St Louis, MO, USA). Mouse anti-α-SMA antibody was purchased from Abcam (Cambridge, MA, USA). The secondary antibodies to each primary antibody were as follows. Anti-mouse IgG HRP conjugated and anti-rabbit IgG HRP conjugated antibodies were obtained from Promega (Madison, WI, USA). Anti-goat IgG HRP conjugated antibody was obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Anti-mouse IgG Alexa flour 488 and anti-rabbit IgG Alexa flour 594 secondary antibodies were purchased from Invitrogen (Carlsbad, CA, USA). Alzet osmotic pump was purchased from Durect corporation (Cupertino, CA, USA).
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