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Author Topic: Blood smear  (Read 10489 times)

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Offline silvia

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Blood smear
« on: January 26, 2009, 11:42:53 AM »
Hello to everyone! I have no experience in ICC  :'( so I'd like to ask you, more expert than me, some easy suggestions.
I set up an antibody against liver and other tissues: now we're interested in using that on a blood smear....
1) Fixation... air-dried or not? If yes... something after? Methanol ethanol or stuff like that?
2) Antigen retrieval... If I use HIER in a tissue for that antbody, is reasonable that the retrieval method could be the same in a blood smear?
Thanks in advance who'll answer me.... stupid questions probably... but not so stupid if it's the first time you perform ICC  ???
Have  a nice day
Silvia

Blood smear
« on: January 26, 2009, 11:42:53 AM »

Offline excalibur

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Re: Blood smear
« Reply #1 on: January 27, 2009, 12:18:25 PM »
Air dry - Yes!

Methanol - Yes!
5 minutes or it will wash off the slide when rehydrated.

Antigen retreival - No! Not needed.
AR is to break formaldehyde bonds to expose the antigen.
Paula Keene Pierce, HTL(ASCP)HT
Excalibur Pathology, Inc.
5830 N Blue Lake Dr.
Norman, OK 73069
405-759-3959
www.excaliburpathology.com

Offline silvia

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Re: Blood smear
« Reply #2 on: January 28, 2009, 05:46:08 AM »
Thanks for reply  :)

Just to check my understainding (not so obvious ;))
1) Air-dry
2) Fix 5 minutes in methanol and after that... go with rehydratation and ICC without ER

Is everything right?

The last question.... are there some epitopes that don't like methanol? Or it can be consider a good universal fixative in ICC?

Thanks again
Silvia

Offline CanuckPhD

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Re: Blood smear
« Reply #3 on: January 28, 2009, 11:21:44 AM »
Yes, some antibodies will not work after methanol fixation. You may have to try acetone. acetone:methanol, PFA etc to find one that works for your antibody.

This fixation comes after the air drying of the smear.

Good luck

Offline silvia

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Re: Blood smear
« Reply #4 on: February 03, 2009, 04:58:05 AM »
Thanks to everyone for your suggestions.
Ciao!  ;)
Silvia

Re: Blood smear
« Reply #4 on: February 03, 2009, 04:58:05 AM »