Anti-Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker (ab487)

  Western blot - Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker (ab487)

All lanes : Anti-Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker (ab487) at 1/1000 dilution

Lane 1 : 0.1% DMSO treated Jurkat cell (whole cell lysate)
Lane 2 : 0.5% DMSO treated Jurkat cell (whole cell lysate)
Lane 3 : 1% DMSO treated Jurkat cell (whole cell lysate)
Lane 4 : 0.1% SDS treated Jurkat cell (whole cell lysate)
Lane 5 : 0.5% SDS treated Jurkat cell (whole cell lysate)
Lane 6 : 1% SDS treated Jurkat cell (whole cell lysate)

Secondary
HRP conjugated goat anti-mouse.
developed using the ECL technique

Performed under reducing conditions.

Observed band size : 84 kDa (why is the actual band size different from the predicted?)


Exposure time : 1 minute

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.

This image is courtesy of an anonymous Abreview

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  Immunocytochemistry/ Immunofluorescence - Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker (ab487)

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.

This image is a courtesy of an anonymous Abreview

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  Immunocytochemistry/ Immunofluorescence-Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker(ab487)

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.

  Flow Cytometry - Nuclear Matrix Protein p84 antibody [5E10] - Nuclear Marker (ab487)

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.