Apoptosis · Antibody review
PUMA
Beyotime · AF1204
After about three years of use, the background remains clean; PUMA decreases over this stimulation time course.
01Experimental images
02Observations
Still performing well after years of use
I have used this PUMA antibody for about three years and remain happy with its performance. The background is clean and the band changes are easy to follow.
A decreasing signal in this experiment
PUMA levels gradually decrease as stimulation continues. By 2 h, accompanying caspase-3 and GSDME measurements show clear activation and cleavage, while PUMA has fallen rather than increased. This is a notable feature of the experiment.
03Protein background
PUMA, encoded by BBC3, is a pro-apoptotic BH3-only member of the BCL2 family, with a common western blot band near 23 kDa. It responds to p53-associated and other stress signals and binds anti-apoptotic proteins such as BCL2 and BCL-XL, helping engage the mitochondrial pathway and cytochrome c release. PUMA expression and turnover vary with cell type, stimulus and time, so different models may show increasing, stable or decreasing protein levels. Conventional western blots primarily track its total abundance.
04Experimental conditions
- Sample
- Human THP-1 monocytes
- Application
- Western blot (WB)
- Primary antibody dilution
- 1:3000
- Gel
- 4–12% gel
- Electrophoresis
- 200 V, 35 min
- Transfer
- 300 mA, 30 min
- Membrane
- 0.2 μm nitrocellulose
- Blocking
- 1% PVP, 1 h
- Secondary antibody
- 1:8000, 1 h
- Protein loading
- Not recorded
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