Effective p21 Antibody, but Optimization Needed

Cell & Tissue Engineering, Imec
Research Scientist

Overall

Quality of Results

Ease-of-Optimization

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Company:

Santa Cruz Biotechnology

Product Name:

Waf1/Cip1/CDKN1A p21 Antibody (F-5)

Catalog Number:

sc-6246

Clone Number:

DNA damage induced by gemcitabine or doxorubicin in bEnd.3 cells was assessed using p21 staining. The same antibody was also evaluated in mouse tissue samples, where target detection proved highly challenging.

Experimental Design and Results Summary

Applications

Immunofluorescence

Sample

bEnd.3 cells, fixed in PFA 4% (10 min @ RT)

Primary Incubation

1 % Triton (10 min @ RT), followed by incubation with primary antibody for 1h @ RT (dilution 1:50 in BSA)

Blocking Agent

1% BSA

Secondary Incubation

AlexaFluor 448 donkey anti-mouse IgG (H+L), dilution 1:800

Tertiary Incubation

none

Detection

Fluorescent microscopy

Results Summary

Successful detection of p21 in bEnd.3 cells following doxorubicin or gemcitabine exposure confirms the induction of DNA damage. However, staining intensity was relatively modest and should always be interpreted in comparison with untreated control cells to distinguish specific signal from background.

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Summary

The Good

Efficient

The Bad

Modest intensity levels. Use of a high dilution. Difficult to detect in IHC.

The Bottom Line

We evaluated several p21 antibodies from Santa Cruz Biotechnology, and this antibody demonstrated the best overall performance.

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