[Histonet] Re IF staining questions

[Amos Brooks]

Hi,
     Direct IiF (a fluorescent conjugated primary) is certainly easier, but
there are some reasons one might prefer using an indirect method (using a
conjugated secondary). Indirect methods allow the use of a different
wavelength to be used simply by switching the secondary. It is also
possible for the fluorescent tag to fade or not be strong enough to start
with. The secondary could be used at a higher concentration or replaced in
the case of a faded primary.

Anos



Message: 1
Date: Thu, 19 Oct 2023 09:22:56 -0400
From: Charles Riley <criley at udel.edu>
To: histonet at lists.utsouthwestern.edu
Subject: [Histonet] IF staining questions
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Why would one decide to use a primary antibody along with a secondary
antibody rather than a primary antibody conjugated with the secondary?

Example.   I have a researcher who wants to do CD11C staining with
Alexafluor488

Is it better to buy and use a primary antibody CD11C conjugated with
Alexafluor488  or to do the CD11C primary and a Rabbit anti-rat (H_L) IgG
antibody secondary?