Isotype-specific immunostaining with mouse primary antibodies on mouse tissue

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.

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.

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.

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 IgG₁heavy chain do stain rat IgG_2aand IgG_2b.)