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Dear all,

thanks so far for your many comments on my problem. I will try to
look through all of these parameters and I am pretty optimistic now I
will find a solution for my problem.

Best wishes to you all,

Florian



>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>Florian,
>
>This is probably precipitated, coagulated, labeled secondary antibody.
>Such precipitates are often present, even in newly purchased
>secondaries. Try preparing your secondary antibody and then filter it
>through a 0.45 um syringe filter before you use it. If you don't have
>syringe filters, then centrifuge your secondary antibody at high G in a
>clinical centrifuge for 5 minutes and then carefully remove the
>supernatant without disturbing any precipitate that might be at the
>bottom of the centrifuge tube and use the supernat for your
>incubations. The syringe filters work best if you have them.
>
>Hope that takes care of the problem,
>
>Steve
>
>____________________________________________________________________
>Stephen C. Kempf
>Associate Professor
>Faculty Director - AU Hybridoma Facility
>Department of Biological Sciences
>131 Cary Hall
>Auburn University, AL  36849
>
>Tel: 334-844-3924
>Fax: 334-844-4065
>
>E-mail: [log in to unmask]
>http://www.auburn.edu/academic/science_math/biology/faculty/kempf.htm
>http://www.auburn.edu/research/hybridoma/
>_____________________________________________________________________
>
>On Wed, 10 Mar 2004, Florian Ulrich wrote:
>
>>  Search the CONFOCAL archive at
>>  http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>  --
>>  dear all,
>>
>>  i am currently doing antibody stainings on paraffin sections.
>>  unfortunately, i am always getting very bright dots when i look at
>>  the staining under the confocal.when i do the control without first
>>  antibody, i also see these dots. therefore, they should clearly come
>>  from the 2nd antibody. it does not matter whether i use an alexa 488
>>  or alexa 546 coupled antibody, they both produce these effects. maybe
>>  the antibodies are bad, but they are from a commercial source and
>>  work fine in many applications. so, is this a probelm related to
>>  paraffin sectioning?
>>
>>  i would appreciate any ideas or experience how to get around that
>>  problem and be forever grateful!
>>
>>
>>  best wishes,
>>
>>  Florian
>>
>>----------------------------------------------------------------------------------------------
>>  Florian Ulrich
>>  Heisenberg Lab
>>  Max-Planck-Institute for Molecular Cell Biology and Genetics
>>  Pfotenhauerstrasse 108
>>  01307 Dresden
>  > Germany
>  > phone: (+49) 351 210 2689
>  > fax:    (+49) 351 210 1489
>  > email: [log in to unmask]
>  >


--


----------------------------------------------------------------------------------------------
Florian Ulrich
Heisenberg Lab
Max-Planck-Institute for Molecular Cell Biology and Genetics
Pfotenhauerstrasse 108
01307 Dresden
Germany
phone: (+49) 351 210 2689
fax:    (+49) 351 210 1489
email: [log in to unmask]