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Western blotAntibody data
- Antibody Data
- Antigen structure
- References [0]
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- Validations
- Western blot [1]
- Immunocytochemistry [1]
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Validation data
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- Product number
- MA1-25100 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- CD146 Monoclonal Antibody (P1H12)
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- Recommended positive controls: human tongue.
- Antibody clone number
- P1H12
- Concentration
- 1.9 mg/mL
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-CD146 Monoclonal Antibody (P1H12) (Product # MA1-25100) and a 130 kDa band corresponding to Cell surface glycoprotein MUC18 was observed across cell lines. Membrane enriched extracts (30 µg lysate) of SK-MEL-31 (Lane 1), SK-BR-3 (Lane 2), HUVEC (Lane 3), HeLa (Lane 4), T-47D (Lane 5), MCF7 (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177,1:4000) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescentdetection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005)..
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Cell surface glycoprotein MUC18 was performed using 70% confluent log phase SK-MEL-28 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with CD146 Monoclonal Antibody (P1H12) (Product # MA1-25100) at 1:100 in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32766), (1:2000), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing Membrane localization. Panel e represents low expression in MCF-7 cells. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
