[Histonet] human macrophages in human melanoma xenografts

Sun Oct 25 16:59:21 CDT 2009

I do not know of a non mouse anti human macrophage marker, but in my
experience most are using the wrong ms monoclonal anti human CD68 for
macrophage markers,  The KP1 clone will stain a lot more than macrophages,
granulocytes etc., the best clone to use for anti human CD68 is the PGM1
clone which is specific to just macrophages and monocytes and not all the
other cells.  I have known hematopathologist who tell me they stopped using
CD68 because it was just not specific enough, but when I tell them to switch
to the PGM1 clone they are very happy with that.

As for macrophages in mouse tissue, I use the serotec rat anti mouse F4/80
ab with very good results.  I have used macrophage markers that are for rat
tissue (can't remember what they are at the moment, I think the are CD68?)
but I did not have as good results on rat tissue with them as I did on ms
tissue with rat anti f4/80.

Liz,
What do you use for macs on rat tissue?  I may have asked you about this
before?

Regards,

Patsy

Patsy Ruegg, HT(ASCP)QIHC
IHCtech
12635 Montview Blvd. Ste.215
Aurora, CO 80045
720-859-4060
fax 720-859-4110
www.ihctech.net 
www.ihcrg.org

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Isaac Milos
Sent: Friday, October 23, 2009 1:57 PM
To: Liz Chlipala; Margaryan, Naira; Galbraith, Joe;
histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts

Liz,

Great point, and definitely something to keep in mind should one do that
stain.  Indeed, CD14 is not specific to macrophages only (monocytes also
express the antigen as discussed).

Of course, there are other more specific macrophage markers (i.e. HAM56)
but these are commonly developed in mice.  The CD14 came to mind as ours
is a rabbit antibody.

Best regards,
Isaac Milos
Cell Marque Corporation

-----Original Message-----
From: Liz Chlipala [mailto:liz <@t> premierlab.com] 
Sent: Friday, October 23, 2009 11:15 AM
To: Isaac Milos; Margaryan, Naira; Galbraith, Joe;
histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts 

She is asking to stain for macrophages and that's a monocyte/macrophage
marker so that's not the same.  You have to be careful with the names of
antibodies and check the specification sheets.  You can search for a
macrophage marker and the antibody may be listed as a macrophage marker
as the following example:

This antibody is from abcam its listed as follows:

Macrophage antibody [MAC387] (ab22506)  but when you look at the
specification sheet it states -- Recognises the L1 or Calprotectin
molecule, an intracytoplasmic antigen comprising of a 12kD alpha chain
and a 14kD beta chain expressed by granulocytes, monocytes and by tissue
macrophages.

So it is not entirely specific to macrophages.  This antibody may work
fine for your application, but if you are looking just to stain
macrophages this is not the antibody you want to pick.

Just a word of advice.

Liz

Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Manager
Premier Laboratory, LLC
PO Box 18592
Boulder, Colorado 80308
office (303) 682-3949 
fax (303) 682-9060
www.premierlab.com

Ship to Address:
1567 Skyway Drive, Unit E
Longmont, Colorado 80504

-----Original Message-----
From: Isaac Milos [mailto:imilos <@t> cellmarque.com] 
Sent: Friday, October 23, 2009 12:01 PM
To: Margaryan, Naira; Liz Chlipala; Galbraith, Joe;
histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts 

Hi Naira,

Our new IVD rabbit monoclonal (anti human) CD14 is preferentially
expressed on monocytes/macrophages:

http://www.cellmarque.com/07/p_detail.php?id=2066 

Best regards,
Isaac Milos
Cell Marque Corp.

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of
Margaryan, Naira
Sent: Friday, October 23, 2009 8:57 AM
To: Liz Chlipala; Galbraith, Joe; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts 

Thanks a lot both of you!

For melanoma detection, I usually use HRP with AEC. Is there any
non-mouse anti-human macrophage marker?

Thank you much,
Naira 

-----Original Message-----
From: Liz Chlipala [mailto:liz <@t> premierlab.com] 
Sent: Friday, October 23, 2009 10:51 AM
To: Galbraith, Joe; Margaryan, Naira; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts 

F4/80 is a mouse macrophage marker.  If she wants to detect human
macrophages in a mouse background she will need to use a mouse
anti-human CD68.  She will need to run it with a mouse on mouse
detection system and run all of the appropriate negative controls.  I
would also select a alkaline phosphatase detection system rather than
HRP with DAB just incase the tumor has any melanin in it.  I have done
this before looking for human lymphocytes with LCA in a mouse
background.  You just need to make sure you have all of the appropriate
controls.  For a positive control I would use a human tonsil or
something like that and you need to make sure you run the appropriate
isotype negative controls.  

Liz

Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Manager
Premier Laboratory, LLC
PO Box 18592
Boulder, Colorado 80308
office (303) 682-3949 
fax (303) 682-9060
www.premierlab.com

Ship to Address:
1567 Skyway Drive, Unit E
Longmont, Colorado 80504

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of
Galbraith, Joe
Sent: Friday, October 23, 2009 9:44 AM
To: Margaryan, Naira; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] human macrophages in human melanoma xenografts 

Naira:

Here is a link to a site listing macrophage markers.  F4/80 is a
commonly mentioned marker for macrophages.  I presume you mean IHC
rather than ICH.  Enjoy.

http://www.antibodybeyond.com/reviews/cell-markers/macrophage-marker.htm

Joe
joseph-galbraith <@t> uiowa.edu

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of
Margaryan, Naira
Sent: Friday, October 23, 2009 10:31 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] human macrophages in human melanoma xenografts 

Hi Histonetters,

I would like to be able to look at human macrophages in human melanoma
xenografts raised in mouse. Could you please suggest me a best Ab for
ICH and protocol?

Thanks in advance,

Naira

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