For electron beam columns, an off-axis alignment assembly for aligning an electron beam emitter tip to an aperture and a method of aligning the two. One embodiment of the alignment assembly is made of an electrically conductive material, such as doped silicon, which is formed integrally with an electron beam microcolumn assembly. The alignment assembly has several areas etched down to thin membranes ranging in thickness from about 2 νm to 10 νm. A target (e.g., a hole or raised area) is formed in each membrane and an aperture is formed during the same operation to maintain precise distances between each of the targeting positions and between each targeting position and aperture. Each membrane is transparent to a light illuminating one, or both, of its surfaces, thus allowing direct alignment of an emitting tip of an electron beam emitter with the target. Once this positioning is performed for, e.g., three targets, the precise location of the emitting tip is known relative to the aperture on the alignemt assembly, thus enabling tip alignment to the aperture without directly viewing either the tip or aperture together.