Optical clearing of murine bones to study megakaryocytes in intact bone marrow using light-sheet fluorescence microscopy

Abstract

In mammals, the differentiation and maturation of megakaryocytes (MKs) occurs in the bone marrow (BM). The three-dimensional environment influences megakaryopoiesis and platelet release. Thus, imaging MKs within the intact BM is important to understand megakaryopoiesis. Here, we present an optical clearing protocol for intact bones and the subsequent microscopic analysis including image processing to quantitatively assess MK size and distribution. This technique overcomes the limitations of classical sectioning methods as the entire bone can be imaged.