Cytoskeleton provides a sheep polyclonal tubulin antibody which recognizes the whole alpha/beta complex of all species from yeast to mammals. It detects methanol fixed (denatured conformation) and paraformaldehyde fixed (native conformation) tubulin in microtubule or the α/β subunit form. See the datasheet below for more information about this antibody.
In addition for staining the actin cytoskeleton, we have developed a unique line of fluorescent phalloidins with improved brightness and stability compared to other conjugates such as Alexa fluor and Cy dyes, for more information see the Acti-stain information page.
Question 1: What is the best way to fix cells for staining with the polyclonal tubulin antibody?
Answer 1: Fix cells with methanol at -20°C for 3 min. . See the ATN02 datasheet for the full protocol.
Question 2: What dilution do you recommend for the polyclonal anti-tubulin antibody?
Answer 2: The dilution of antibody depends on the cell/tissue type and experimental format being used. For western blotting, we have tested and recommend using the antibody at a concentration of 500 ng/ml (1:1000 dilution). At this dilution, we detected tubulin in extracts of Drosophila S2 cells, Xenopus A6 cells, mouse Swiss 3T3 cells, rat NRK cells, human HeLa cells, and bovine brain. For immunofluorescence, we used the anti-tubulin antibody to detect tubulin in mouse Swiss 3T3 cells. The antibody was used at a concentration of 2.5 μg/ml (1:200 dilution) followed by incubation with a 1:500 dilution of anti-sheep rhodamine-conjugated secondary antibody. This information is found usually in the figure legends or within the “Product Uses” section of our antibody datasheets.
For more information, click on the Document tab above to see the datasheet.