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Author Topic: cytospin fixation and storage  (Read 19890 times)

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Offline braz

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cytospin fixation and storage
« on: March 03, 2006, 11:21:42 AM »
I need to store some unstained cytospin slides and I've found the most divergent opinions on how to store it. After lifting the cells out of the culture flasks, I've been ressuspending them in Cytospin Collection Fluid to spin. Should I fix them in 95% ethanol before storing? What should be the ideal temperature to store? Some people say room temperature, others say -80...

cytospin fixation and storage
« on: March 03, 2006, 11:21:42 AM »

Offline ImmunoNYC

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Re: cytospin fixation and storage
« Reply #1 on: March 04, 2006, 02:25:21 PM »
Whether or not you want to fix and what you want to fix with will solely depend on what sort of antigen you are staining and which antibody you will be using.

To keep my options open I routinely let them dry a few hours - or sometimes overnight - at RT after removing from the cytospin machine.  If I am not planning to do staining immediately, I will store them, wrapped well in a slide box with dessicant, at -80 deg C to preserve antigenicity until I need them. When I need them (note that this is for my antigen/antibody of interest) I remove them from -80 deg C and directly immerse in acetone. I have also had to use methanol and PFA depending on what I am looking for.

I know a lot of people fix in acetone BEFORE storing at -80 deg C. This should be fine too but what happens if your antigen will only work on formaldehyde fixed cells? That is the problem I have encountered and is the explanation for why I fix after freezing.

The major drawback of fixing after freezing is that you can get freezing artifacts if you let lots of nasty ice crystals build up on your slides (usually from opening and closing a freezer or slide box multiple times without protecting your slides). Unless you store them properly in dry conditions my method won't consistently give you good results.

Hope this helps! :)
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Offline braz

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cytospin fixation and storage
« Reply #2 on: March 06, 2006, 01:48:48 AM »
Thanks. That really helped!

cytospin fixation and storage
« Reply #2 on: March 06, 2006, 01:48:48 AM »