RE: double IHC/in situ

From:Robert Geske <>

here is what i found


-----Original Message-----
From: Gayle Callis []
Sent: Friday, January 12, 2001 2:10 AM
To: Marek Malecki;
Subject: Re: double IHC/in situ

What is  website address, please. 

 At 04:55 AM 1/11/01 -0800, you wrote:
>Dr. Gwen Childs wrote the whole chapter on the subject in 1995: Specimen 
>Preparation for Microscopy and Microanalysis.  SM International Press, 
>Chicago, IL.
>In addition, her www site is loaded with protocols and images.
>> >Hello Histonetters,
>> >
>> >Some of the researchers around here do in situ hybridization followed
>> >by IHC.  However, the harsh conditions of the in situ hybridization
>> >techniques destroys many of the antigenic sites of interest.  Does
>> >anyone out there use antigen retrieval to recover antigenicity
>> >following in situ techniques, or does anyone know of any references?
>> >
>> >Karen Larison
>> >University of Oregon
>>Hi Karen,
>>ISH techniques sometimes work like "antigen retrieval techniques" for some
>>antigens. I have used ISH in combination with ICC for pituitary hormones,
>>and the ICC combined to the ISH was often stronger and with less
>>than alone. However, for other antigens, the ISH procedures seem a bit too
>>harsh, and no antigen retrieval will recover this.
>>We've tried to combine ISH with ICC for vasopressin, and the immunolabel
>>was strongly reduced. I suspect that for small peptides like vasopressin,
>>the proteinase K step combined with the high temperatures of the ISH
>>in elutin of some of the peptide.
>>We have tried microwave antigen retrieval for vasopressin and other
>>antigens like intermediate filaments, Pit-1, thyroid hormone receptors and
>>several other antigens, but nothing would do for any of those. I think you
>>will have to try, but I suspect that antigen retrieval will be of little
>>help after in situ.
>>                           -=-
>>                          (o -) O
>>Paul Klosen, PhD
>>CNRS UMR 7518 Neurobiologie des Fonctions Rythmiques et Saisonnieres
>>Universite Louis Pasteur
>>12 rue de l'Universite
>>F-67000 Strasbourg, FRANCE
>>tel:    fax:
>Marek Malecki, M.D., Ph.D.
>Director and Principal Investigator
>Molecular Imaging Laboratories, University of California, San Diego
>address: 1500 Suite Bonner Hall, 9500 Gilman Drive, La Jolla, CA92093-0368
>phone: 8588223373
>fax: 8584573355
>pager: 8586161420
>www site:
>ftp site:
Gayle Callis
Veterinary Molecular Biology
Montana State University
Bozeman MT 59717-3610
406 994-4705
406 994-4303

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