Some suggestions for starting out in IHC and keeping it cheap are:

Get the DAKO Handbook in IHC and use it as your reference

If at all possible, fix samples in zinc formalin for short time to illiminate
most antigen retrieval

Buy a semi-universal detection system which will work with at least mouse and
rabbit primary antibodies, and goat if goat primaries are anticipated, this way
you won't have to worry about matching up your detection with your primary

Start with primary antibodies that are tried and true, so that pretreatments and
dilutions are spelled out

Use concentrated antibodies to save money

Humid chambers and manual processing served us all well for many years.  Cheap
processors which are not automated include Shandon's slides attached to chamber
wells (?Sequenza) and I think Fisher still sells the manual capillar gap
staining system first designed by Bragatti

Good controls will be your best friend.  If you can find them, tissues both
positive and negative for the antigen of interest are important to run.
Negative controls using the isotype of your primary antibody, i.e. (if your
primary antibody is mouse IgG1A, then your negative control should be mouse
IgG1A at the same protein concentration dilution and time as your primary).
Another neg. control is BSA or buffer in place of the primary antibody to check
for quenching of endogenous peroxidase, biatin, and or non specific binding of
your detection reagents

Make sure you dry your sections completly onto silane coated slides so they
don't fall off

If you are a member of NSH you can join the Immunohistochemical Resource Group
by going to our web site at: fill out the form and mail it in
This is a group of nearly 300 people from all over the world working in IHC, and
we share our expertise with each other.  We have an email listserv which
connects us all by email, a reference list for IHC, tissue and reagent bank for
controls and donated reagents for IHC, and on and on

Good luck, and we are only an email away.

Patsy Ruegg

"Ryan, Genoula K Ms USAMH" wrote:

> To HistoNetters,
> Our new Chief of Pathology would like us to do a few immunos.  We are not
> set up to do any.  Is there a cheap way to do immunos?  We don't want to
> purchase a Ventanna or anything fancy.  Our workload is small and he only
> wants the basics (i.e., S100, CK, ER/PR).  Any suggestions on how to get
> this up and running will be greatly appreciated.  Also if there are any good
> reference books for IHC.  Thanks in advance,
> Genie K. Ryan,
> Histology Supervisor
> DSN: 371-2653
> 49-6221-172653

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