Alexa Fluor® 488 Anti-p53 antibody [SP5]

3 product images

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  Flow Cytometry - Alexa Fluor® 488 Anti-p53 antibody [SP5] (ab270192)

  Flow cytometry overlay histogram showing MCF7 cells stained with ab270192 (red line). The cells were fixed with 4 % formaldehyde (10 min) and then permeabilized with 0.1 % PBS-Triton X-100 for 15 min. The cells were incubated in 1x PBS containing 10 % normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab270192) (1x 10⁶ in 100µl at 0.2 µg/ml (1/2500)) for 30 min at 22°C.

  Isotype control antibody (black line) was Rabbit IgG (monoclonal) Alexa Fluor 488 ^® (Alexa Fluor® 488 Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab199091) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

  Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.

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  Immunocytochemistry/ Immunofluorescence - Alexa Fluor® 488 Anti-p53 antibody [SP5] (ab270192)

  Immunofluorescence staining of p53 in A431 cells using ab270192. The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab270192 at 1/100 dilution (shown in green) and Alexa Fluor® 647 Anti-Tubulin antibody [YOL1/34] - Microtubule Marker ab195884, Rat monoclonal to Tubulin (Alexa Fluor^® 647), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
  

  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Alexa Fluor® 488 Anti-p53 antibody [SP5] (ab270192)

  IHC image of p53 staining in a section of formalin-fixed paraffin-embedded human colon cancer*.

  The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6) in a Biocare Medical NxGen pressure cooker using retrieval settings of 110°C for 20 minutes. Non-specific protein-protein interactions were then blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M (w/v) glycine and 1% (w/v) BSA for 1h at room temperature. The section was then incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab270192 at 1/100 dilution (shown in green) and counterstained using Alexa Fluor® 647 Anti-Tubulin antibody [YOL1/34] - Microtubule Marker ab195884, Rat monoclonal to Tubulin (Alexa Fluor^® 647), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Fluoromount^®.

  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.

  *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.