Rapid, simple, column-free isolation of CD11b+ cells from various mouse organs (INC6P.323)

Abstract

Abstract CD11b+ cells are distributed throughout the body and act as sentinels for the immune system. The frequency of CD11b+ myeloid cells varies in different organs, ranging from approximately 48 % of the brain myeloid (microglia) cells, 30% of bone marrow and 6% of total nucleated cells in gut and spleen. The integrin CD11b heterodimerizes with CD18 to form the mature complex Mac-1. Mac-1 facilitates a variety of immune cell responses including phagocytosis, adhesion, migration and chemotaxis, all functions that are crucial for normal tissue homeostasis and inflammatory responses. CD11b+ cells are difficult and time consuming to isolate, due to their variable frequency using traditional methods. Here, we describe EasySepTM, a simple, column-free immunomagnetic method for the positive selection of highly purified CD11b+ cells from brain, bone marrow, spleen and gut. Labelled CD11b+ cells are retained in a tube in an EasySepTM magnet while unwanted cells are poured off. Selection can be fully automated using RoboSepTM. Starting with a frequency of 7.6 ± 1.3 % CD11b+ cells in spleen, purities of 93.9 ± 4.5 % (n=41) were achieved. Fully competent cells, as shown by functional assays (phagocytosis, ROS), were isolated from the above tissues. This EasySepTM technology provides a fast and simple method to access highly purified functional CD11b+ cells from a variety of tissues, facilitating their study ex-vivo.