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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [3]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
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- Product number
- MA5-26812 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- PHGDH Monoclonal Antibody (OTI5C4)
- Antibody type
- Monoclonal
- Antigen
- Recombinant full-length protein
- Reactivity
- Human, Mouse, Rat
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- OTI5C4
- Vial size
- 100 µL
- Concentration
- 1 mg/mL
- Storage
- -20° C, Avoid Freeze/Thaw Cycles
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of PHGDH in HEK293T cells in untransfected (Left lane) and transfected (Right lane) samples using 5 µg per lane. The samples were separated by SDS-PAGE and probed with PHGDH (Product # MA5-26812) monoclonal antibody.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of PHGDH in HEK293T cells in untransfected (Left lane) and transfected (Right lane) samples using 5 µg per lane. The samples were separated by SDS-PAGE and probed with PHGDH (Product # MA5-26812) monoclonal antibody.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot was performed using Anti-PHGDH Monoclonal Antibody (OTI5C4) (Product # MA5-26812) and a 57 kDa band corresponding to D-3-phosphoglycerate dehydrogenase was observed across cell lines and tissue tested, except A549, MDA-MB-231, and SK-O-V3 which shows low to negative expression for PHGDH as reported in the literature. Whole-cell extracts (30 µg lysate) of NTERA-2 cl.D1 (Lane 1), A549 (Lane 2), HCT 116 (Lane 3), MDA-MB-231 (Lane 4), HeLa (Lane 5), LNCaP (Lane 6), SK-O-V3 (Lane 7), Raji (Lane 8), Mouse Brain (Lane 9) and Rat Brain (Lane 10) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 (1:2000) 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 PHGDH was performed using 70% confluent log phase NTERA-2 cl.D1 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 PHGDH Monoclonal Antibody (OTI5C4) (Product # MA5-26812) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then with Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32723) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with Hoechst 33342 (Product # H1399). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 40X magnification in CellInsight CX7 LZR High-Content Screening (HCS) Platform (Product # CX7C1115LZR).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry was performed on paraffin-embedded human liver tissue. To expose target proteins, 1 mM EDTA in 10mM Tris, pH8.5, 120°C for 3min was used. Following antigen retrieval, tissues were probed with a PHGDH monoclonal antibody (Product # MA5-26812).
