acetocarmine staining of microspores

Paul A Bucciaglia bucc0003 at
Wed Apr 6 00:51:24 EST 1994

Hello folks,

I've been staining tobacco microspores (about 3-5 days post tetrad) with
acetocarmine to follow microspore mitosis.  Using the std. 1% carmine
in 45% acetic acid on both fixed (ethanol/acetic acid) and unfixed antehrs
gives me bright red cytoplasms that obscure any sign of a nucleus.  i dug
up a reference from 1937 (Maheshwari, Stain Tech. 12#2) which suggested
a few drops of chloral hydrate to clear the cytoplasm.  "ahh, an archaic
name for HCl" i thought so i added a drop of 1N HCl.  This did help some
as the nucleus was just barely visible and the cytoplasm cleared somewhat.
So i checked the Sigma catolog and was surprised to find that chloral 
hydrate is a nasty chlorinated hydrocarbon that is (was?) used to make DDT.

So my question:  is there a (much) less toxic compound to clear the micro-
spore cytoplasm and allow me to visualize the nucleus?

please note i have tried to heat the slides after squasshing out the 
micropsores. this helps some if followed by a drop of 1N HCl but I would
like to see more intense staining/less background.

Thanks for bearing with me,

paul bucciaglia


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