Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [1]
- Immunocytochemistry [2]
- Flow cytometry [2]
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Validation data
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- Product number
- 433140 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- PODXL Monoclonal Antibody
- Antibody type
- Monoclonal
- Antigen
- Other
- Reactivity
- Human
- Host
- Mouse
- Isotype
- IgM
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- -20°C
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-PODXL Monoclonal Antibody (Product # 433140) and a 160 kDa band corresponding to PODXL was observed in all cell lysates tested except Hep G2 and A549. Whole cell lysates (30 µg lysate) of Hep G2 (Lane 1), NCCIT (Lane 2) and A549 (Lane 3) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris 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 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection 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 GCTM-2 was performed using 70% confluent log phase HEK 293 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with PODXL Mouse Monoclonal Antibody (Product # 433140) at 2µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L)/IgM (L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing membranous localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of PODXL was performed using 70% confluent log phase NCCIT and A549 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 PODXL Monoclonal Antibody (Product # 433140) at 2 µg/mL 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) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing localization to plasma membrane. Panel e shows A549 cells with no expression of PODXL. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Staining of BG01V Cells Provided by Jon Chesnut (Invitrogen), shows a dose dependant immunostaning.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Staining of BG01V Cells Provided by Jon Chesnut (Invitrogen), shows a dose dependant immunostaning.