The localization of B lymphocytes in mouse bone marrow: radioautographic studies after in vivo perfusion of radiolabelled anti-IgM antibody

An in vivo cell surface labelling technique using radioautography has been developed to visualise the distribution of IgM-bearing B lymphocytes within the bone marrow. Anaesthetized 3-week-old mice were perfused via the common iliac artery with: (1) serum-containing medium (SCM), (2) 125I-labelled anti-IgM antibody in SCM, (3) SCM, and (4) fixative. In radioautographic sections of femoral marrow labelled surface IgM+ cells were observed either singly or in small clusters throughout the extravascular haemopoietic marrow cords. Binding specificity was demonstrated by the displacement of 125I-anti-IgM labelling by excess anti-IgM and by the binding of perfused 125I-anti-H-2Kk antibody in CBA/J (H-2Kk) mice but not in C57BL/6 (H-2Kb) mice. Quantitative analysis of radioautographic sections revealed an even distribution of labelled cells throughout CBA/J marrow perfused with 125I-anti-H-2Kk, indicating a uniform accessibility of perfused antibody to cells in the haemopoietic cords. This labelling pattern contrasted with that in 125I-anti-IgM perfused animals in which surface IgM+ cells, although widely distributed in the bone marrow, showed areas of concentration, speculatively clones of maturing B lymphocytes. This method of labelling surface IgM and other cell markers in situ provides an approach to study the microenvironment of B lymphocyte genesis in the bone marrow.