Hi everyone.

My cultivated tumor granulosa cells are undergoing a senescence-inducing treatment.

At the end of it, I am counting SA-beta-gal+ cells, so I want to counterstain nuclei with hematoxylin, as follows:

1) washing cells with 1X PBS and with filtered tap water for 1 minute;

2) adding 250 μL of hematoxylin solution (according to Mayer) [Sigma-Aldrich, cat. no. 51275] for 5 minutes;

3) washing cells twice filtered tap water for 30 seconds;

4) adding filtered tap water and observing under the inverted microscope.

Beta-gal positive cells are easily identifiable, but nuclear counterstaining is very weak, especially at high magnifications.

Can anyone help me find a way to have a more pronounced signal from the hematoxylin staining, please?

Thanks a lot in advance.

PS I already tried longer staining times.

Stefano