Determine the primary antibody details including the host species in which it was produced, its sensitivity, validation, specificity to your target antigen, and compatibility with your immunoassays.
Select a host species of the primary antibody that is different from the host species in which your samples were collected.
Choose a secondary antibody that will bind to the primary antibody. The secondary antibody that you choose must have been produced in a host species that is different from the host species in which the primary antibody was produced. For example, you will need an anti-mouse secondary antibody for a mouse primary antibody. Both antibodies must be produced in different host species.
The secondary antibody that you choose must match the class or subclass of the primary antibody. For example, choose an anti-mouse IgM secondary antibody when using a primary monoclonal antibody from the mouse IgM class. An IgG secondary antibody is a better option if you don’t know the immunoglobulin classes or subclasses of the primary antibody.