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Author Topic: Bone marrow cytospins fixation  (Read 8994 times)

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Offline cepaiva

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Bone marrow cytospins fixation
« on: February 12, 2006, 10:22:55 AM »
Is my first time in this forum and I would like to have some informations. I am trying to search for isolated tumor cells in the bone marrow aspirates of breast cancer patients. We will use real time-PCR and immunocytochemistry. My questions:
1. What type of fixative is best in my case (we'll use MetCheck Kit AE1/AE3 with secondary antibody-AP)?
2. Can I frozen the glasses (-80) before fixation?

Bone marrow cytospins fixation
« on: February 12, 2006, 10:22:55 AM »

Offline excalibur

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Bone marrow cytospins fixation
« Reply #1 on: February 12, 2006, 12:07:37 PM »
The protocol should be included with the MetCheck kit.

Ask Dako what fixative they recommend if it is not.

Methanol, acetone, formalin, could all be used.

I would airdry the slides, fix them, and then store if you are not able to stain right away.
Paula Keene Pierce, HTL(ASCP)HT
Excalibur Pathology, Inc.
5830 N Blue Lake Dr.
Norman, OK 73069
405-759-3959
www.excaliburpathology.com

Offline cepaiva

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bone marrow cytospins fixative
« Reply #2 on: February 12, 2006, 05:40:40 PM »
Thanks Paula.
And what about the store the glasses...do you think we can store them at -80? Is it harmful for the glasses?Do I have to put the cytospins in a special box to be frozen?

Thanks
Carlos

Offline cepaiva

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bone marrow cytospins fixative
« Reply #3 on: February 12, 2006, 05:47:21 PM »
Another consideration, Paula.

I don't know if you understand...I'm colecting my materials yet but I don't have the kit yet, so I need to prepare the materials (fixe and frozen) before to receive the Kit. In the Dako's kit the fixative is a mix of formalin and alchool (but I don't know the exact proportions). Do you think it's harmful to fix them with another fixative (acetone or formalin for example)?

Thanks again
Carlos

Offline excalibur

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Bone marrow cytospins fixation
« Reply #4 on: February 12, 2006, 06:50:01 PM »
Hello Carlos,

yes, I thought you might not have received the kit yet because Dako is very good about including protocols. AE1/AE3 is a very good antibody and should work well no matter what fixative. I recommend it should contain some formalin. A 20% alcohol/80% formalin should work nicely.

When doing just iron stains, we always fix the smears in 100% methanol for ~ 5 mins.

Call Dako tech support first thing Mon. morning and ask them what fixative they recommend.

You can air dry the slides and store in the freezer until you are sure what the recommended fixative is.
Paula Keene Pierce, HTL(ASCP)HT
Excalibur Pathology, Inc.
5830 N Blue Lake Dr.
Norman, OK 73069
405-759-3959
www.excaliburpathology.com

Bone marrow cytospins fixation
« Reply #4 on: February 12, 2006, 06:50:01 PM »