Anti-Nuclear Matrix蛋白p84抗体[5E10] (ab487)

7.5% SDS-PAGE gel.

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4% paraformaldehyde-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained for Nuclear Matrix Protein p84 (green) using ab487 at 1/500 dilution in ICC/IF.

Red: phalloidin, a cytoskeleton marker, at 1/200 dilution.

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SDS and DMSO were constituents of the lysis buffer. 0.1% SDS did not break down the nucleus entirely , however the higher concentrations did and p84 was detected in the lysate.

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ab487 staining Nuclear Matrix Protein p84 in Human stomach adenocarcinoma cell line (AGS) by Immunocytochemistry/ Immunofluorescence. The cells were formaldehyde fixed, permeabilised in 0.025% Triton X-100, TBS and then blocked using 5% serum for 1 hour at 23°C. Samples were then incubated with primary antibody at 2µg/ml for 1 hour at 23°C. The secondary antibody used was a goat anti-mouse IgG conjugated to Alexa Fluor® 350 (blue) used undiluted.p84 shows nuclear localization.

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ICC/IF image of ab487 stained Hepp 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 (ab487, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 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.

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Overlay histogram showing HeLa cells stained with ab487 (red line). The cells were fixed with 100% 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 (ab487, 1µg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

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Nuclear Matrix Protein p84 was immunoprecipitated from HepG2 (human liver hepatocellular carcinoma cell line) whole cell lysate with 3 µg ab487. Western blot was performed from the immunoprecipitate using ab487. Anti-Rabbit IgG was used as a secondary reagent.

Lane 1: HepG2 whole cell lysate 30 μg.

Lane 2: Control IP in HepG2 whole cell lysate with 3 μg of pre-immune mouse IgG.

Lane 3: ab487 IP in HepG2 whole cell lysate.

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7.5% SDS-PAGE gel.

This image was generated using the ascites version of the product.

Acetone-fixed frozen section of human stomach tissue stained for Nuclear Matrix Protein p84 using ab487 at 2 µg/ml in immunohistochemical analysis.

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