RE: cell culture for IHC

From:Luis Chiriboga

Hi All
I have seen a system that allows you to grow cells directly on glass slides
.  I do not know the name of  the system since I have never actually used
for cell culture but do have a collaborator who use it exclusively.
Essentially,  It is a clear plastic box without bottom that fits over the
glass slide (excluding the label area).  A "rubber?" contact adhesive is
used(do not know name or composition) to attach the box to the slide surface
such that a water tight seal is made. The box comes with a lid and the cells
can be grown directly on the glass slide. The benefits of this system are
that any fixative can be used.  Also,  if you are doing manual staining,
the adhesive acts as a hydrophobic barrier so you do not have to use a PAP
pen.  The main drawback is if your are doing automated staining.  You have
to scrape the adhesive off the slide for cap gap instruments or ventana
instruments because it will impeded the movement/mixing of solutions.  In
addition,  the entire slide is covered with cells so it would require a lot
of reagent when doing manual staining, we have gone around this by wiping of
areas of the slide Ill see if I can come up with a name.
Thought this might be another option (although the lack of vendor
information  makes it this post quite useless :-)

-----Original Message-----
From: Chris van der Loos []
Sent: Friday, February 14, 2003 2:53 AM
Subject: RE: cell culture for IHC

Hi Laura,
In the past I did some IHC on cell cultures grown on plastic disposables.
This created an extra problem with respect to the fixation since
acetone-fixation was not possible for obvious reasons. Finally, we used a
4% paraformaldehyde (freshly prepared in PBS) for 5 min at RT. This short
aldehyde-fixation does not cause any cross-linking and therefore antigen
retrieval is not needed (impossible too, because of the plastics!)
During ALL IHC incubation steps and washings (from endo PO blocking up to
chromogen) we included 0.1% saponin to open up the cell membranes (to get
your IHC reagents in and out of the cells).
Lots of success!

Chris van der Loos
Dept. of Cardiovascular Pathology
Academic Medical Center
Amsterdam, Netherlands

Laura wrote:
 >Date: 13 Feb 2003 10:30:26 -0600
 >Subject: Cell Culture for IHC
 >I have been asked to do an immunohistochemistry stain (West Nile Virus)
on cell
 >culture. I usually on worked with paraffin blocks and smears. I'm not even
 >sure of the questions to ask, but here it goes....
 >In growing the cells, can I used glass slides (I don't have any plastic)?
 >Are the glass slides plain or charged?
 >How do I fix the cells? (I know if they're not fixed in formalin there
 >be no antigen retrieval.)
 >Anything special I need to know before the jump such as potential
problems or
 >background staining?
 >Thanks in advance,
 >Laura Bliven
 >Marshfield Laboratories

Kathleen wrote:
 >Date: 13 Feb 2003 13:01:22 -0600
 >From: Kathleen Spencer 
 >Subject: Re: Cell Culture for IHC
 >In the past I did IHC on cultured hippocampal neurons grown on round
 >glass coverslips in tiny petri dishes. The coverslips were coated with
 >poly-l-lysine after soaking in nitric acid. Everything was done with
 >sterile technique of course. The fixation of the cells and the staining
 >took place in the petri dish, suctioning off each solution. The
 >coverslips were placed cell side down on a slide with vectashield for
 >Hope this helps,


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