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Synthetic peptide derived from residues 500 - 600 of Human NSD3.
Our Abpromise guarantee covers the use of ab4514 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use a concentration of 5 µg/ml.|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 72.6, 161 kDa (predicted molecular weight: 72.6 , 161 kDa).|
|IHC-P||1/50 - 1/100. Perform heat mediated antigen retrieval via the microwave method before commencing with IHC staining protocol.|
Breast carcinoma tissue was fixed in 10% formalin at 4°C overnight and paraffin embedded. Following blocking the tissue sections were incubated with ab4514 (1/50 dilution) at 37°C for 1 hour, and with the secondary antibody at 37°C for 30 minutes. The slides were counterstained with Hematoxylin. Washes were carried out using TBS. Heat mediated antigen retrieval was carried out using the Microwave method before commencing the staining protocol. ab4514 staining appears to be localized mainly to the cytoplasm. Although the putative cellular localization of NSD3 is the nucleus we have found no evidence in the current literature to confirm this and we believe ab4514 may be detecting a cytoplasmic form of the protein. We would be very grateful for any customer feedback about NSD3 and ab4514.
ICC/IF image of ab4514 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab4514, 5µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
As seen in Immunohistochemical analysis (shown above), although the putative cellular localization of NSD3 is the nucleus we have found no evidence in the current literature to confirm this and we believe ab4514 may be detecting a cytoplasmic form of the protein.