RE: [Histonet] Universal (protein) block for IHC

From:"Hermina Borgerink"

I routinely use casein in both my diluent as well as wash buffer for all
my immunos (I primarily use an alkaline phosphatase detection system,
but when doing double or triple immuno labeling I also use an HRP or
Beta Galactosidase system).  Before converting from the standard
protocol which used a non-immune serum from the same species as the
primary for blocking non-specific proteins, I ran a comparison and found
there to be no difference at all.  I use a Tris wash buffer that
contains 0.5% casein as well as 0.1% Tween 20 as a surfactant (I use the
MicroProbe system which employs capillary action).  My slides are clean
and have virtually no background staining.  The only time when I have
noticed some non-specific binding is when using goat primaries.  If
anyone would like my protocol, please respond privately since I cannot
attach on Histonet.


Hermina M. Borgerink, BA, HTL(ASCP)QIHC
Wake Forest University Health Sciences
Department of Pathology
Medical Center Blvd.
Winston-Salem, NC 27157
Tel. (336) 716-1538
Fax. (336) 716-1515
"Treat a man as he appears to be, and you make him worse.  But treat a
man as if he already were what he potentially could be, and you make him
what he should be."  (Goethe)
This electronic message, including any attachments, is confidential and
proprietary and is solely for the intended recipient.  If you are not
the intended recipient, this message was sent to you in error and you
are hereby advised that any review, disclosure, copying, distribution or
use of this message, or any of the information included therein, is
unauthorized and strictly prohibited.  If you have received this
electronic transmission in error, please immediately notify the sender
by reply and permanently delete all copies of this message and its
attachments.  Thank you. 

-----Original Message-----
[] On Behalf Of Philip
Sent: Thursday, April 07, 2005 11:01 AM
Subject: RE: [Histonet] Universal (protein) block for IHC


Casein is the main protein in milk. Skim milk has lots more stuff in 
it than just casein, and who knows what that's doing to your sections?


>I thought casein was just a fancy name for skim milk.  I use it
>sometimes as an avidin block by making a 5% solution out of cheap dried
>-----Original Message-----
>[] On Behalf Of Kelly D
>Sent: Wednesday, April 06, 2005 4:42 PM
>To: Johnson, Teri
>Cc: Histonet
>Subject: Re: [Histonet] Universal (protein) block for IHC
>Hi Teri,
>I noticed a decrease in specific staining when I plugged it into my
>protocol. I made it myself and it was a big pain. Buy it! Try
>antibody concentration on a few slides and see what happens. I never
>a block with my secondary.
>Good luck!
>Johnson, Teri wrote:
>>Has anybody converted to using a universal protein block (casein) for
>>their immunohistochemistry protocols rather than using species
>>normal sera?  For those who have made the conversion, did you have to
>>make any changes to your methodology or did you just plug it in to
>>protocol and run?  Did you notice any difference in the staining
>>intensity for known antibodies (stronger, weaker, no change)?  Are you
>>also using it as a secondary antibody diluent or for washes?  Finally,
>>are you using commercially available reagent or are you making your
>>Thanks so much!
>>Teri Johnson
>>Managing Director Histology Facility
>>Stowers Institute for Medical Research
>>1000 E. 50th St.
>>Kansas City, Missouri  64110
>>Histonet mailing list
>Histonet mailing list
>Histonet mailing list

Philip Oshel
Supervisor, BBPIC microscopy facility
Department of Animal Sciences
University of Wisconsin
1675 Observatory Drive
Madison,  WI  53706
voice: (608) 263-4162
fax: (608) 262-5157 (dept. fax)

Histonet mailing list

Histonet mailing list

<< Previous Message | Next Message >>