Staining mouse tissue with anti-mouse Ig (H&L chain)
secondary reagents inevitabily causes staining of interstitial immunoglobulins,
B cells, plasma cells and macrophages with Ig bound to Fc receptors.
To avoid this, staining with F(ab) monomeric secondary
has been developed (References). Several
companies sell kits to do this and Jackson
Immunoresearch Laboratories Inc sells all the components to develop
in-house such staining.
A cheap alternative is to exploit the relative lower levels
of each IgG isotype in serum and on follicular B cells to stain with isotype-specific,
conjugated, anti-mouse secondary antibodies. IgG isotypes account each
for 20-25% ot total serum Ig. In addition, young laboratory mice raised
in sterile, pathogen free barriers and unimmunized have lower endogenous
immunoglobulin levels than older animals raised in conventional housing.
Most mouse monoclonal antibodies working on mouse tissue
are of IgG1 isotype.
Detailed explanation of this method can be found in:
Cattoretti G, Qing Fei. Application of the Antigen Retrieval
Technique in experimental Pathology: from human to mouse. In Antigen
Retrieval Techniques. Shi SR, Gy J and Taylor CR Editors. Eaton Publishing,
Natick MA 2000. pp. 165-179.
(Can be obtained by Biotechnique
Press. A limited number of reprints are also available upon request:
e-mail)
Briefly, substitute your anti H+L pan-reactive secondary antibody with an anti-isotype heavy chain specific secondary. Be aware of possible odd crossreactivties (e.g. many goat anti mouse IgG1 heavy chain do stain rat IgG2a and IgG2b.)