For immunofluorescent staining of the aortic tissue sections, we used sodium citrate (0.01 M, pH 6.0) for antigen retrieval. Sections were then blocked for 30 min with 5% BSA in PBS at room temperature. The following diluted primary antibodies: mouse anti-BKCa α (1:200, Abcam, Cambridge, MA, UK), rabbit anti-BKCa β (1:1000, Abcam, Cambridge, MA, UK), rabbit anti-Kir6.1 (1:200, Santa Cruz, CA, USA), mouse anti-Kir6.2 (1:50, Santa Cruz, CA, USA), rabbit anti-Cav1.2 (1:100, Santa Cruz, CA, USA), goat anti-Cav1.3 (1:200, Santa Cruz, CA, USA), were used to incubate sections overnight at 4 °C. Slides were then incubated at 37 °C with secondary antibodies Alexa Fluor 488 (donkey anti-goat, 1:400; Life Technologies), Alexa Fluor 488 (donkey anti-rabbit, 1:400; Life Technologies), Alexa Fluor 488 (donkey anti-mouse, 1:400; Life Technologies), Alexa Fluor 594 (donkey anti-rabbit, 1:400; Life Technologies), Alexa Fluor 594 (donkey anti-mouse, 1:400; Life Technologies) and Alexa Fluor 594 (donkey anti-goat, 1:400; Life Technologies) for 1 h. 4′,6-diamidino-2-phenylindole (DAPI) (blue) was used to counterstain all sections. The slides were then mounted and viewed under a fluorescent microscope (Olympus BX53).
Rabbit anti cav 1
Manufactured by Santa Cruz Biotechnology
Sourced in United States
The Rabbit anti-Cav-1 is an antibody that recognizes the Caveolin-1 (Cav-1) protein. Cav-1 is a structural component of caveolae, which are invaginations of the plasma membrane that play a role in various cellular processes. The Rabbit anti-Cav-1 antibody can be used to detect and study the expression and localization of Cav-1 in biological samples.
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Lab products found in correlation
Mouse anti bkca α, by Abcam (2 mentions)
Rabbit anti bkca β, by Abcam (2 mentions)
Rabbit anti kir6, by Santa Cruz Biotechnology (2 mentions)
Mouse anti kir6, by Santa Cruz Biotechnology (2 mentions)
Goat anti cav1, by Santa Cruz Biotechnology (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 594, by Thermo Fisher Scientific (1 mentions)
Dm4000b microscope, by Leica (1 mentions)
Diaminobenzidine tetrahydrochloride dab, by Merck Group (1 mentions)
Image pro plus 6, by Media Cybernetics (1 mentions)
Rabbit anti egfr, by Santa Cruz Biotechnology (1 mentions)
Mouse anti mhc, by Santa Cruz Biotechnology (1 mentions)
Mouse anti α tubulin, by Merck Group (1 mentions)
Rabbit anti cavin1, by Abcam (1 mentions)
Mouse anti cav3, by BD (1 mentions)
Ventana bench mark xt autostainer, by Roche (1 mentions)
5 protocols using rabbit anti cav 1
1
Immunofluorescent Staining of Aortic Tissues
2
Immunohistochemical Analysis of Ion Channels
3
Immunohistochemical Analysis of Colorectal Cancer
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Immunostaining was performed on 5-μm formalin-fixed, paraffin-embedded tissue sections using an immunoperoxidase method with rabbit anti-Cav-1 (1: 400; Santa Cruz, Dallas, Texas, USA); rabbit anti-EGFR (1: 100; Santa Cruz, Dallas, Texas, USA), and rabbit anti-Ki-67 (1: 100; Sunbiote, Shanghai, China) monoclonal antibodies. Protein was visualized using the PV and DAB chromogenic kits (Beijing Zhongshan Golden Bridge Biotechnology) following the manufacturer’s instructions.
Two pathologists blinded to the experiments assessed the extent and intensity of immunostaining. The protein expression levels of Cav-1 and EGFR were observed under high-power magnification (×400); 5 different fields of view were randomly selected in each section, and 200 cells were counted. The degree of staining was scored between 0 and 3 points: 0 point=no staining, 1 point=weak staining, 2 points=moderate staining, and 3 points=strong staining. Cells scoring >2 points were defined as positive cells; <50% of positive cells within a field was defined as a negative expression, and ≥50% of positive cells was defined as a positive expression. We chose EGFR-positive expression tissues for the subsequent study. A total of 76 cases of colorectal cancer tissues were further tested for the expressions of Cav-1 and Ki-67.
Two pathologists blinded to the experiments assessed the extent and intensity of immunostaining. The protein expression levels of Cav-1 and EGFR were observed under high-power magnification (×400); 5 different fields of view were randomly selected in each section, and 200 cells were counted. The degree of staining was scored between 0 and 3 points: 0 point=no staining, 1 point=weak staining, 2 points=moderate staining, and 3 points=strong staining. Cells scoring >2 points were defined as positive cells; <50% of positive cells within a field was defined as a negative expression, and ≥50% of positive cells was defined as a positive expression. We chose EGFR-positive expression tissues for the subsequent study. A total of 76 cases of colorectal cancer tissues were further tested for the expressions of Cav-1 and Ki-67.
4
Cavin-1, Caveolins, and ERK Signaling
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The following primary antibodies were used: rabbit anti-Cavin-1 (AbCam, Cambridge, UK, 1:3000 dilution) for immunoblotting and immunofluorescence analyses; rabbit anti-Cavin-1 (ThermoScientific, Waltham, MA, USA) for immunoprecipitation assay; rabbit anti-Cav-1 (Santa Cruz Biotechnology, Dallas, TX, USA, 1:1000 dilution); mouse anti-Cav-1 (Zymed-Life Technologies, Monza, Italy, 1:100 dilution); mouse anti-MHC (Santa Cruz Biotechnology, 1:1000 dilution); mouse anti-Cav-2 (BD, Buccinasco, Italy, 1:1000 dilution); mouse anti-Cav-3 (BD, 1:1000 dilution); mouse anti-total and -phosphorylated ERK1/2 (Tyr204) (Santa Cruz Biotechnology, 1:1000 dilution); mouse antialpha-tubulin (Sigma-Aldrich, 1:10 000 dilution).
5
Immunohistochemical Analysis of Cav1 Expression
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Cells were grown on sterilized coverslips (22 mm × 22 mm) in Petri dishes and washed in phosphate-buffered saline (PBS 1×) for 5 min and fixed in 4% neutral-buffered formalin for 10 min.
Xenograft and tissue samples were fixed in 4% buffered formaldehyde and paraffin-embedded. Threemicrometer-thick sections from the mouse tissue samples as well as from representative tumor blocks of the human adenocarcinoma samples were collected on Superfrost Plus slides for immunohistochemistry.
Cav1 immunohistochemistry was performed using an automated slide-processing platform (Ventana BenchMark XT AutoStainer, Ventana Medical Systems, Tucson, AZ, USA) and a polyclonal antibody (rabbit anti-Cav1, diluted 1:350; Santa Cruz Biotechnology). Cav1 staining was assessed as a categorical variable (negative or positive if present in at least 10% of neoplastic cells) and analyzed independently by three of the authors (R.S., E.D., and P.C.). Vascular endothelium represented an internal positive control. In discrepant cases, slides were reviewed at a multiheaded microscope to reach agreement.
Xenograft and tissue samples were fixed in 4% buffered formaldehyde and paraffin-embedded. Threemicrometer-thick sections from the mouse tissue samples as well as from representative tumor blocks of the human adenocarcinoma samples were collected on Superfrost Plus slides for immunohistochemistry.
Cav1 immunohistochemistry was performed using an automated slide-processing platform (Ventana BenchMark XT AutoStainer, Ventana Medical Systems, Tucson, AZ, USA) and a polyclonal antibody (rabbit anti-Cav1, diluted 1:350; Santa Cruz Biotechnology). Cav1 staining was assessed as a categorical variable (negative or positive if present in at least 10% of neoplastic cells) and analyzed independently by three of the authors (R.S., E.D., and P.C.). Vascular endothelium represented an internal positive control. In discrepant cases, slides were reviewed at a multiheaded microscope to reach agreement.
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