pollen maturity status determination - (May/03/2006 )

Hi everyone!
I asked around in my lab about a procedure to determinate pollen maturity status. They found for me this protocol to stain pollen and see if it's mononucleate or binucleate. Unfortunately no one here has never done it. This is the protocol:

Take the flower and immerse it overnight in Farmer's solution (3:1 absolute ethanol/acetic acid + 1.9% FeCl3 x 4 H2O)
Cut anthers and smash them on a microscope glass slide
Add some drops of acetocarmine 2% solution
Cover the specimen and observe

Now I've got plenty of doubts. These are the most urgent ones:

1. I have to apply this protocol on closed flowers. Do I have to use entire flower? Won't it be a problem to remove anthers from inside of the calyptra after overnight bath?
2. Do I have to wash acetocarmine after staining and before observing the slide under the microscope?
3. Is this really the best way to check anther maturity?

Hey Ila,
It will be better if you take flower buds of different size (fresh) smear them in 2% acetocarmin slon with a iron needle or scalepel put coverslip and observe under microscope. I hope you will sure get result. We never kept flower buds overnight in any fixative.
Beena