The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 - 5 µg/ml. Detects a band of approximately 39 kDa (predicted molecular weight: 39 kDa).
Use at an assay dependent concentration. Detection limit is approximately 3ng/ml as a capture antibody when used against the immunogen.
Use 0.5µg for 106 cells. ab170191-Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Use a concentration of 5 µg/ml.
FunctionSingle-stranded nucleic acid binding protein that binds preferentially to oligo dC. Major cellular poly(rC)-binding protein. Binds also poly(rU). Negatively regulates cellular antiviral responses mediated by MAVS signaling. It acts as an adapter between MAVS and the E3 ubiquitin ligase ITCH, therefore triggering MAVS ubiquitinationa and degradation.
Tissue specificityDetected in all tissues examined.
Sequence similaritiesContains 3 KH domains.
Post-translational modificationsPhosphorylated. The non-phosphorylated form(s) exhibited the strongest poly(rC)-binding activity.
Cellular localizationNucleus. Cytoplasm. Loosely bound in the nucleus. May shuttle between the nucleus and the cytoplasm.
ICC/IF image of ab77323 stained MCF7 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab77323, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96879, DyLight® 488 goat anti-mouse IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
IHC image of PCBP2/hnRNP E2 staining in Human normal testis formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab77323, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Western blot - PCBP2/hnRNP E2 antibody (ab77323)
Anti-PCBP2/hnRNP E2 antibody (ab77323) at 5 µg/ml + K562 cell lysate at 50 µg
Secondary Goat Anti-Mouse IgG (H&L)-HRP Conjugate at 1/5000 dilution Developed using the ECL technique
Predicted band size : 39 kDa Observed band size : 39 kDa Additional bands at : 35 kDa. We are unsure as to the identity of these extra bands.
Overlay histogram showing HeLa cells stained with ab77323 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab77323, 0.5µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 1µg/1x106 cells) used under the same conditions. Unlabelled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
References for Anti-PCBP2/hnRNP E2 antibody (ab77323)
This product has been referenced in:
Saul MJ et al. UPF1 regulates myeloid cell functions and S100A9 expression by the hnRNP E2/miRNA-328 balance. Sci Rep6:31995 (2016).
Read more (PubMed: 27573788) »