merosin and emerin on frozen muscle sections.

From:manal galal

Dear colleagues,
             My work is on frozen muscle. I am having trouble with merosin and emerin immunostaining on frozen muscle. I wonder if my freezing process is to blame? 
    My freezing procedure is as follows:
1-We recieve the specimen immediately after the biopsy is taken.
2-We put it in a fridge for 15 min to relax.
3- The section is stuck on a pre-frozen chuck with a dab of OCT.
4- It is then immediately placed on the quick freeze shelf (-40C) of the cryostat for 15 min.
5-Then the specimen is put in the main compartment of the cryostat        (-20C) and left there for 20 min. to accomodate to the temperature.
6- The sections are then cut at 8 microns and placed on the slides.
7- The slides are left to dry overnight at room temperature.
By the way we get very good results with Dystrophins and Sarcoglycans, using this method of freezing.
Any suggestions??
Thank you
 Manal Galal

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