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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [1]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
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- Product number
- MA1-847 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- RANBP2 Monoclonal Antibody (2E1)
- Antibody type
- Monoclonal
- Antigen
- Recombinant full-length protein
- Description
- MA1-847 detects Nup358/RanBP2 from human samples.
- Reactivity
- Human
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- 2E1
- Vial size
- 100 µg
- 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 Nup358 was performed by loading 25 µg of cytoplasmic and nuclear HeLa cell lysates prepared using NE-PER Nuclear and Cytoplasmic Extraction Reagents (Product # 78833) per well, 16 µg of HeLa whole cell lysate prepared using M-PER Mammalian Protein Extraction Reagent (Product # 78501), and 10 µL of PageRuler Plus Prestained Protein Ladder (Product # 26619) onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288), and blocked with StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) for 1 hour at room temperature. Nuclear-enriched Nup358 was detected above 460 kDa after probing with a Nup358 monoclonal antibody (Product # MA1-847) at a dilution of 1:1000 in blocking buffer overnight at 4C on a rocking platform, washing in TBST, and probing with an HRP-conjugated goat anti-mouse IgG secondary antibody (Product # 31430) at a dilution of 1:20,000 for 1 hour. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34076).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of Nup358 (green) in HeLa cells. Cells were fixed with 4% paraformaldehyde, permeabilized with 0.1% Triton X-100 in PBS, and blocked with 1% Blocker BSA in PBS (Product # 37525), each for 15 minutes at room temperature. Cells were stained with a Nup358 monoclonal antibody (Product # MA1-847) at a concentration of 10 µg/mL in 1% Blocker BSA in PBS, followed by a DyLight 488-conjugated goat anti-mouse IgG secondary antibody (Product # 35502) at a dilution of 1:1000 for 1 hour at room temperature. Nuclei (blue) were stained with DAPI (Product # 46190). Images were taken on a Thermo Scientific ToxInsight Instrument at 20X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry was performed on normal biopsies of deparaffinized human stomach tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a Mouse Monoclonal Antibody recognizing Nup358 (Product # MA1-847) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.
