Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

A bleaching/quenching step should solve the problem.


I.  IF THE TISSUE IS PARAFFIN EMBEDDED, FIRST REHYDRATE.

   1.      Xylene --> 100% alcohol --> 100% alcohol --> 95% alcohol -->
70% alcohol --> 50% alcohol  --> 30% alcohol --> water
   2.      PBS wash

II.  REDUCE BACKGROUND

   1.      Incubate with NaIO4 (21.4 mg/ml in PBS) 15 min. followed by PBS
wash
   2.      Alternative bleaching solutions can be fresh NaBH4 or glycine
followed by PBS wash

III.  BLOCK WITH NORMAL SERUM INCUBATION  (etc.)

from
http://www.aecom.yu.edu/aif/instructions/immunofluor/protocol01.htm


> ----- Original Message -----
> From: "Soumitra Ghoshroy" <[log in to unmask]>
> To: <[log in to unmask]>
> Sent: Friday, October 06, 2006 4:12 PM
> Subject: brain autofluorescence
>
>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> Hello Everyone,
>>
>> We have an user who wants to label an enzyme in hypothalamus using
>> immunoflurescence. We used Alexa Fluor 647 as secondary in paraffin
>> embedded
>> tissue and discovered that the tissue is autofluorescing in almost every
>> laser
>> line. I am not at all familiar with brain tissue.  We also used
>> unlabeled
>> deparaffinized sections and looked at it and the autofluorescence level
>> is
>> extremely high. So I would like to get some advice on how to image it.
>> Is
>> there
>> anything we are not doing right? The tissue was fixed with 4%
>> paraformaldehyde.
>>
>> Thanks in advance.
>>
>> Soumitra
>>
>>
>> *******************************************************************************
>> Soumitra Ghoshroy
>> College Associate Professor, Biology
>> Director, Electron Microscopy Lab
>> New Mexico State University
>> Las Cruces, NM 88003
>> Tel: 505-646-3268 (office)
>>     505-646-3283 (lab)
>> Fax: 505-646-3282
>> http://biology-web.nmsu.edu/ghoshroy/ghoshroy.htm
>> http://emldata.nmsu.edu
>>
>>
>


_________________________________________
Michael Cammer
Analytical Imaging Facility and
Dept. ASB Biophotonics Innovation Laboratory
Albert Einstein College of Medicine
1300 Morris Park Avenue, Bronx, NY  10461
718-430-2890  Fax 718-430-8996
work:  http://www.aecom.yu.edu/aif/
personal:  http://coxcammer.com/