E2F1 Recombinant Rabbit Monoclonal Antibody [JJ092-02]

Overview

Product Name

E2F1 Recombinant Rabbit Monoclonal Antibody [JJ092-02] - BSA and Azide free

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within Human E2F1 aa 371-404 / 437.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IF-Cell, IF-Tissue, IHC-P, IP, FC

Molecular Weight

Predicted band size: 47 kDa

Positive Control

HT-29 cell lysate, SW480 cell lysate, SW620 cell lysate, HepG2 cell lysate, NIH/3T3 cell lysate, rat heart tissue lysate, Hela, MCF-7, HepG2, human tonsil tissue, mouse pancreas tissue, rat heart tissue.

Conjugation

unconjugated

Clone Number

JJ092-02

Product Features

Form

Liquid

Concentration

1mg/ml

Storage Instructions

Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles.

Storage Buffer

1*PBS (pH7.4).

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

WB
1:1,000-1:5,000

IF-Cell
1:100-1:500

IF-Tissue
1:100-1:500

IHC-P
1:1,000-1:5,000

FC
1:1,000

IP
1-2μg/sample

Target

Function

The human retinoblastoma gene product appears to play an important role in the negative regulation of cell proliferation. Functional inactivation of Rb can be mediated either through mutation or as a consequence of interaction with DNA tumor virus-encoded proteins. Of all the Rb associations described to date, the identification of a complex between Rb and the transcription factor E2F most directly implicates Rb in regulation of cell proliferation. E2F was originally identified through its role in transcriptional activation of the adenovirus E2 promoter. Sequences homologous to the E2F binding site have been found upstream of a number of genes that encode proteins with putative functions in the G1 and S phases of the cell cycle. E2F-1 is a member of a broader family of transcription regulators including E2F-2, E2F-3, E2F-4, E2F-5, E2F-6 and E2F-7 each of which forms heterodimers with a second protein, DP-1, forming an "active" E2F transcriptional regulatory complex.

Background References

1. Peng W & Feng J Long noncoding RNA LUNAR1 associates with cell proliferation and predicts a poor prognosis in diffuse large B-cell lymphoma. Biomed Pharmacother 77:65-71 (2016).

2. Millour J et al. ATM and p53 Regulate FOXM1 Expression via E2F in Breast Cancer Epirubicin Treatment and Resistance. Mol Cancer Ther 10:1046-58 (2011).

Sequence Similarity

Belongs to the E2F/DP family.

Post-translational Modification

Phosphorylated by CDK2 and cyclin A-CDK2 in the S-phase. Phosphorylation at Ser-364 by CHEK2 stabilizes E2F1 upon DNA damage and regulates its effect on transcription and apoptosis.; Acetylation stimulates DNA-binding. Enhanced under stress conditions such as DNA damage and inhibited by retinoblastoma protein RB1. Regulated by KAP1/TRIM28 which recruits HDAC1 to E2F1 resulting in deacetylation. Acetylated by P/CAF/KAT2B.

Subcellular Location

Nucleus.

UNIPROT

SwissProt: Q01094 Human

SwissProt: Q61501 Mouse

SwissProt: O09139 Rat

Synonyms

DmelCG6376 antibody

Dmel_CG6376 antibody

drosE2F1 antibody

E(Sev-CycE)3A antibody

E(var)3-93E antibody

E2-promoter binding facto antibody

E2F 1 antibody

E2F transcription factor 1 antibody

E2F-1 antibody

E2f-PA antibody

Expand

Images

Western blot analysis of E2F1 on different lysates with Rabbit anti-E2F1 antibody (HA750322) at 1/500 dilution.

Lane 1: HT-29 cell lysate
Lane 2: SW480 cell lysate
Lane 3: SW620 cell lysate
Lane 4: HepG2 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 47 kDa
Observed band size: 60 kDa

Exposure time: 3 minutes; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA750322) at 1/500 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
Western blot analysis of E2F1 on different lysates with Rabbit anti-E2F1 antibody (HA750322) at 1/1,000 dilution.

Lane 1: NIH/3T3 cell lysate (10 µg/Lane)
Lane 2: Rat heart tissue lysate (20 µg/Lane)

Predicted band size: 47 kDa
Observed band size: 60 kDa

Exposure time: 1 minute 2 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA750322) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
ICC staining of E2F1 in Hela cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (HA750322, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ICC staining of E2F1 in MCF-7 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (HA750322, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ICC staining of E2F1 in HepG2 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (HA750322, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-E2F1 antibody (HA750322) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA750322) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded mouse pancreas tissue with Rabbit anti-E2F1 antibody (HA750322) at 1/5,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA750322) at 1/5,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded rat heart tissue with Rabbit anti-E2F1 antibody (HA750322) at 1/5,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA750322) at 1/5,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Flow cytometric analysis of HepG2 cells labeling E2F1.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA750322, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
Flow cytometric analysis of NIH/3T3 cells labeling E2F1.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA750322, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
E2F1 was immunoprecipitated from 0.2 mg HepG2 cell lysate with HA750322 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA750322 at 1/1,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: HepG2 cell lysate (input)
Lane 2: HA750322 IP in HepG2 cell lysate
Lane 3: Rabbit IgG instead of HA750322 in HepG2 cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 26 seconds; ECL: K1801

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"