Problem with microtome sectioning - (Apr/23/2008 )

hi guys
I don't know if this is the right place to ask this question
but since I can't find anyone else to ask
so I will give it a try

I am cutting parafilm embede mouse lung into 4um, it used to be 5um
but the section came out too thick
however the most terrible question I am facing
is that the septum of the PBS (control) group seems extended
it should be like the one circled by black
the septum circled by red is the places that the septum looks extended

[attachment=4585:PBS.jpg]
I did not pull the ribon while cutting
and I did not hesitated during the cuttig

Can someone help me out?

thanks

cutting is probably not the problem - fixation or dehydration may be
i'm afraid sometimes you just have to put up with this stuff
have you considered frozen tissue and a cryostat as a true control (dont fix it just slap on some eosin and check the morphology)

good luck

dom

you cant avoid it - you can only alter it
everything you do to the tissue changes it in some way (otherwise you wouldnt bother doing it) - the trick is to use stuff which alters your region of interest as little as possible
dehydration by alcohol will create shrinkage as the water is removed
fixation reduces the effect of antibodies on your epitopes
are you just staining or antibody labelling - a confocal microscope would mean you could cut thicker sections (less ripping)

dom