Disappearing Cytoplasm!!

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From:Bruce Abaloz <b.abaloz@zoology.unimelb.edu.au>
Date:Mon, 20 Sep 1999 17:22:03 +1000
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Hi all....I'm a student here @ the Uni.of Melbourne & am freezing fresh liver & spleen (mouse), floating the OCT covered specimen in a boat in liquid nitrogen
until OCT is a solid block then I drop it all into the liquid nitrogen. I store it @ -70C until ready to cut in the cryostat.I am using Histogrip slides & cut @ 6 microns @ -10C.I let it dry @ room temp.for 1/2 hr.- 2 hrs.then fix in 10% NBF for 10 min. rinsing X3 in PBS. after which I transfer to DH2O to transport to the Histology Lab. for H&E staining. I have no problem with the stain but the sections after staining, look as if the cytoplasm has been "dehydrated away".Can
anyone advise me on what "to do" or what I'm doing wrong. I've also tried Acetone fixation (10 min.@ RT) with the same result??????????? I look forward to your replies & THANKYOU in advance.........Simone Cuff.


DEPARTMENT of ZOOLOGY	         *  ph:    +61 3 93446282
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Parkville Victoria.3052          *  email: b.abaloz@zoology.unimelb.edu.au                                                                       
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