The mammary gland is a highly dynamic tissue that changes throughout reproductive life, including growth during puberty and repetitive cycles of pregnancy and involution. Mammary gland tumors represent the most common cancer diagnosed in women worldwide. Studying the regulatory mechanisms of mammary gland development is essential for understanding how dysregulation can lead to breast cancer initiation and progression. Three-dimensional (3D) mammary organoids offer many exciting possibilities for the study of tissue development and breast cancer. In the present protocol derived from Sumbal et al., we describe a straightforward 3D organoid system for the study of lactation and involution ex vivo. We use primary and passaged mouse mammary organoids stimulated with fibroblast growth factor 2

The mammalian neocortex, the outer layer of the cerebrum and most recently evolved brain region, is characterized by its unique areal and laminar organization. Distinct cortical layers and areas can be identified by the protein expression of graded transcription factors and molecular determinants that define the identity of different projection neurons. Thus, specific detection and visualization of protein expression is crucial for assessing the identity of neocortical neurons and, more broadly, for understanding early and late developmental mechanisms and function of this complex system. Several immunostaining/immunofluorescence methods exist to detect protein expression. Published protocols vary with regard to subtle details, which may impact the final outcome of the immunofluorescence.