[Histonet] Re: Reduction of autofluorescence using glycine

[Bob Nienhuis]

If this works by binding free aldehyde groups that attach to antibodies/ or
fluorochromes,  or biotinylated whatever. shouldn't it also work for DAB or
ABC immunolabeling and
reduce background labeling?

Bob
UCLA / VA Medical Center

On Fri, Dec 12, 2008 at 2:08 PM, Gayle Callis <gayle.callis <@t> bresnan.net>wrote:

> To reduce aldehyde induced autofluorescence, you can use 100 - 300 mM
> glycine in pH 7.4 buffer.  TRIS buffer or even Dulbeccos PBS will work.  You
> rehydrate the section and then immerse into the glycine solution for 20
> minutes, maybe even longer.  Glycine works by getting rid (binding?) of free
> aldehyde groups.  You can either treat the tissue prior to processing (after
> fixation) by immersing for an hour or so, but we simply did the glycine
> treatment on individual sections.  It worked best for us when we did a short
> length fixation in NBF.
>
> This has been discussed at length on Histonet in the past, so do an archive
> search.  One person put a summary together on various methods and what
> worked best for him.
>
> There are other methods for getting rid of autofluorescence although some
> are less successful than others and one is made from a chemical that is
> explosive.   Try IHCworld website, fluorescence topics  or Google access
> this discussion written by Wright Cell Imaging Faculty, Toronto Western
> Research Institute, titled:  Autofluorescence, Causes and Cures, a must read
> on the subject.
>
> Another trick is to use fluorophores in the near infrared range, the camera
> sees the fluorescence but no autofluorescence and you cannot see this red
> fluorophore with the naked eye.  Alexa 750 will work if you have the filters
> and excitation wavelength available.
>
> Good luck
>
> Gayle M. Callis
> HTL(ASCP)HT,MT
> Bozeman MT
>
>
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