Magnesium AZ31B alloy has been subjected to uniaxial and pure shear testing inside a scanning electron microscope to elucidate the nature of plastic deformation and its initiation and growth using real-time electron backscatter diffraction (EBSD) lattice orientation analysis. A novel anti-buckling guide was developed to allow for EBSD during uniaxial compression testing and a new method for preparing magnesium samples for analysis using a 3D-printed vibrational polishing machine was developed. The initial formation of twins and their subsequent growth and evolution during sample testing was studied to ascertain their crystallographic type, distribution and alignment relative to different stress and strain states. Transmission electron microscopy (TEM) was used to obtain further details of twins such as initiation sites within grains and twin-precipitate interaction.