MA3-070

antibody from Invitrogen Antibodies

Targeting: KPNB1 IMB1, Impnb, IPO1, IPOB, MGC2155, MGC2156, MGC2157, NTF97

Provider product page for MA3-070

Western blot

Immunocytochemistry

Immunoprecipitation

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [28]
Comments [0]
Validations
Immunocytochemistry [3]
Flow cytometry [3]
Other assay [3]

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Product number: MA3-070 - Provider product page
Provider: Invitrogen Antibodies
Product name: Importin beta-1 Monoclonal Antibody (3E9)
Antibody type: Monoclonal
Antigen: Purifed from natural sources
Description: MA3-070 detects Importin beta 1 from human, canine, bovine, rat and mouse tissues as well as porcine cell lysates. Importin beta 1 is also known as NTF97, Nuclear Transport Factor p97, and karyopherin beta 1. This antibody does not detect bacterially expressed recombinant NTF97. MA3-070 has been successfully used in Western blot, immunofluorescence, and immunoprecipitation procedures. By Western blot, this antibody detects a single ~97 kDa protein representing NTF97 in MDBK cell homogenate. Immunofluorescence staining of NTF97 in MDBK cells with MA3-070 results in the labeling of both the nuclear envelope (NE) and the cytoplasm and shows discontinuous, punctate staining along the rim of the NE. This antibody can inhibit import of nuclear localization sequence-containing proteins in permeabilized cells. The MA3-070 antigen is purified NTF97 from bovine erythrocytes.
Reactivity: Human, Mouse, Rat, Bovine, Canine, Porcine
Host: Mouse
Isotype: IgG
Antibody clone number: 3E9
Vial size: 100 µg
Concentration: 1 mg/mL
Storage: -20° C, Avoid Freeze/Thaw Cycles

KPNB1 protein structure - MA3-070 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Importin beta-1 in Jurkat cells (green) compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde and washed with PBS. Cells were blocked with a 2% solution of BSA-PBS for 30 min at room temperature and incubated with a Importin beta-1 monoclonal antibody (Product # MA3-070) at a dilution of 1 µg/test for 40 min at room temperature. Cells were then incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated secondary antibody and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Importin beta-1 in NIH-3T3 cells (green) compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde and washed with PBS. Cells were blocked with a 2% solution of BSA-PBS for 30 min at room temperature and incubated with a Importin beta-1 monoclonal antibody (Product # MA3-070) at a dilution of 1 µg/test for 40 min at room temperature. Cells were then incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated secondary antibody and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Importin beta-1 in PC12 cells (green) compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde and washed with PBS. Cells were blocked with a 2% solution of BSA-PBS for 30 min at room temperature and incubated with a Importin beta-1 monoclonal antibody (Product # MA3-070) at a dilution of 1 µg/test for 40 min at room temperature. Cells were then incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated secondary antibody and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: RNA immunoprecipitation (RIP) western of KPNB1 was performed on K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 µg of KPNB1 monoclonal antibody (Product # MA3-070) rotating 60 min at RT. The immune complexes were captured on 625 µg of anti-mouse coated Dynabeads (Product # 11202D), washed extensively, and eluted with NuPAGE™ LDS Sample Buffer (Product # NP0007). Samples were resolved onto NuPAGE™ 4-12% Bis-Tris gel (Product # NP0335BOX). Lanes 1 and 3 are input and lanes 2 and 4 are IP. Proteins were transferred to PVDF membrane (Product # IB23001). Membrane was blocked in 5% milk. Target was detected using a KPNB1 monoclonal antibody (Product # MA3-070) at a dilution of 1:2000, followed by a 1:4000 dilution of secondary antibody. Chemiluminescent detection was performed using ECL Western Blotting Substrate (Product # 32106). Data courtesy of the Yeo lab as part of the ENCODE project (www.encodeproject.org).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 3. Identification of nuclear import receptor for NFAT5. (A) Quantitative analysis of subcellular localization of FLAG fusion protein fluorescence signal in HeLa cells transfected with FLAG-NFAT5 132-264 PEPCK and siRNA targeting the indicated members of the Kapbeta family, followed by treatment with isotonic (Iso) or hypertonic (Hyper) medium for 90 min. Cells were stained with FLAG antibody and FITC-labeled secondary antibody, then counterstained with DAPI. At least 100 cells were scored in each condition. Data are presented as mean+-s.e.m. of three independent experiments. (B) Western blotting analysis showing cytoplasmic (C) and nuclear (N) distribution of endogenous NFAT5 in HeLa cells transfected with siRNA targeting KPNB1 under different extracellular tonicities. NF-90 and alpha-tubulin antibodies were used as nuclear and cytoplasmic markers, respectively. Blots are representative of three independent experiments. (C) Left: co-immunoprecipitation analysis of KPNB1 and NFAT5. Immunoprecipitation was carried out using anti-KPNB1 antibodies, and immunocomplexes were subjected to western blot analysis using the NFAT5 and KPNB1 antibodies. Crude input lysate (5% total lysate) and IgG-only samples are shown as controls. Right: co-immunoprecipitation analysis of FLAG-NFAT5 132-581 and KPNB1. Immunoprecipitation (IP) was carried out using anti-FLAG affinity resin, and immunocomplexes were subjected to western blot analysis using the KPNB1 and FLAG antibodies. Blots shown a