Western blotting Focal adhesion Kinase (FAK)

Western blotting is a widely used technique to size separate proteins from a pool of cell or tissue lysates. The technique has 4 major steps: a) gel electrophoresis, b) blocking and treatment with antigen specific antibody, c) treatment with secondary antibody and finally d) detection and visualization. Though western blotting is a widely used technique, detection of specific proteins depends on several factors, the major ones are antibody concentration, incubation time and washing steps. Key points for obtaining clean blots are: always prepare fresh buffer solutions and optimize antibody concentration. Given the advent of high-throughput protein analysis and a push to limit the use of lab consumables, onestep antibodies are developed which recognise protein of interest and also contain a detection label.

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Found 3 matching solutions for this experiment

Upstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.
Protocol tips
-Rabbit (1:500)
-To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for
5 min. Lysates can be aliquoted and stored at -20°C for future use.
Upstream tips
-Store as concentrated solution. Centrifuge briefly prior to opening vial.For short-termstorage(1-2 weeks), storeat 4ºC.For
long-term storage, aliquot and storeat -20ºC or below. Avoid multiple freeze-thaw cycles.
Protocol tips
-Rabbit (1:1000)
Protocol tips
-Rabbit (1:1000)
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