[Histonet] overprocessed plant tissue?

From:Julien LambreyDeSouza

Hello,

I am trying to cut embedded white Birch embryos. Because the seedcoat 
(mainly sclérenchyme) is too hard, we take the embryos out of the seedcoat=20
and process them.

I think, we're getting overprocessed (fried?) embryos, but then again, I am=20
unexperienced with plant tissues. What would I have to look for in order to=20
determine overprocessed tissue?
The cells in the embryo are clearly visible, but their contents seems to 
have binded into spherical masses, like small pink marbles (colored in 
eosine). The nucleus sometimes appears blue (heamatox) in between the masses.

The processing goes as follows:
EtOH 70% 30min
EtOH 80% 30min
EtOH 90% 30min
EtOH100% 30min
EtOH100% 30min
EtOH100% 30min
EtOH100% 30min
EtOH100% + Xylene (50/50) 30min
Xylene 30min
Xylene 45min
Paraffin 45 min
Paraffin 1H30

Coloration:
Xylene 3min
Xylene 3min
Xylene 3min
EtOH 100% 2min
EtOH 100% 2min
EtOH 100% 3min
EtOH 95% 2min
EtOH 95% 2min
EtOH 70% 2min
Water 2min
Hematoxylin 4min
Water 2min
Bluing reagent 2min
water 3min
EtOH 95% 30sec
Eosine 2min
EtOH 95% 1min
EtOH 95% 1min
EtOH 100% 1min
EtOH 100% 2min
EtOH 100% 3min
Xylene 3min
Xylene 3min

I posted some pics to the histonet site this morning. They should be their=20
in 24H. Look for "plant embryo-10x.jpg", "plant embryo-40x.jpg" (3 pics) 
and "plant embryo-100x.jpg".
In the meantime, does anybody have any suggestion to what the problem may be?

Thanks,
Julien Lambrey de Souza
Biologie Évolutive,
Université du Québec à Rimouski
(418) 723-1986 #1714 


_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet


<< Previous Message | Next Message >>