Exploring the Role of Microbiota in the Limiting of B1 and MZ B-Cell Numbers by Naturally Secreted Immunoglobulins.

Immunoglobulins (Igs)-or antibodies (Ab)-are important to combat foreign pathogens but also to the immune system homeostasis. We developed the AID-/- μS-/- mouse model devoid of total soluble Igs and suitable to monitor the role of Igs on immune homeostasis. We used this experimental system to uncover a negative feedback control of marginal zone (MZ) and B1 B cells numbers by naturally secreted Igs. We raised AID-/- μS-/- mice in germ-free conditions demonstrating that this effect of natural secreted Igs is independent of the microbiota. Herein, we provide a comprehensive description of the protocols to establish and use the AID-/- μS-/- mice to study the role of total secreted Igs or of different Ig classes. This study involves Igs injections to AID-/- μS-/- mice or establishment of AID-/- μS-/- mixed bone marrow chimeras that provide a powerful system to study AID-/- μS-/- B cells in the presence of stable concentrations of different Ig classes. While we describe flow cytometric and histological methods to analyze MZ and B1 B cell subsets, AID-/- μS-/- mice can be used to study the effects of natural Igs on other B cell subsets or immune cells.