Abcam
Anti-Survivin Antibody
ab24479
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Immunohistochemistry (IHC) on formalin/PFA-fixed paraffin-embedded sections (FFPE)
Human tissue sections (pancreas)
Heat mediated antigen retrieval. Buffer/Enzyme Used: DAKO antigen retrieval buffer, pH 9.0; Permeabilization in Triton 0.1% in TBS 10min; Antibody dilution 1/50 in TBS, incubated 1 hour at 37°C
1% BSA, incubated 1 hour at 37°C
Anti-rabbit/mouse IgG supplied in the DAKO Envision HRP Kit (Cat No. K4065 from Dako, an Agilent Technologies company)
N/A
DAKO Envision HRP Kit according to manufacturer’s instructions; counterstaining with hematoxylin, microscopic detection and validation.
I could not get reliable results. I had difficulties reproducing results (background variations, staining variations). I attempted to optimize by changing staining conditions such as time, temperature or antibody concentration. I also tried staining smears of pancreatic cell lines, but again could not obtain consistent results. This antibody seems to have some affinity to meiotic cells, but even then, it generates staining with high background.
After de-paraffinization, antigen retrieval and permeabilization, slides were treated for 10min with 0.25% H2O2 to block endogenous phospatases. After incubation with the primary Ab, slides were washed 3x 10min in TBS-Tween.
Acceptable for yes/no experiments, sensitivity is very high.
Difficulties in reproducing results, specificity is questionable.
At least with pancreatic staining, specificity is too low for rating expression patterns.