Latest on Cyclin D1 immunostaining

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From:Keith Miller <> (by way of histonet)
To:histonet <>
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Dear Cyclin D1 sufferers,

For several years we employed 1mM EDTA pH8.0 retrieval for cyclin D1, but
have recently changed to the Dako pH9.9 Target Retrieval Solution.
Microwave Oven heat in 600ml of this solution, starting from cold, for 25
minutes,  and flush out with cold running water at the end of the heating
cycle. Alternatively, the fluid can be saved and re-used, by  placing the
hot container in cold running water for 10 minutes and then promptly
transferring the slides into the cold water.

Incubate in Dako clone DCS-6 at 1/100 for one hour and label with
avidin-biotin px etc. [DCS-6 is also available from many other suppliers]

The pH9.9 fluid is much more efficient than pH8.0 EDTA and has made the
demonstration of Cyclin D1 in mantle cell lymphoma as reliable as using
cytokeratin antibodies for labelling epithelium.

However, I would not recommend using this high pH retrieval solution in a
pressure cooker as sections can sometimes detach.

I hope this helps.

Unfortunately, I no longer subscribe to the Histonet on a regular basis, so
if further details are required, please feel to contact me directly.

Keith Miller
Royal Free & University College London Medical School

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