Longitudinal shear strength and shear modulus of spruce and larch wood with a maximum of micro- and macro-structural variability was determined using a new testing method. Oven-dry density and slope of grain were measured after the shear tests. For the spruce wood samples, a data set of fiber and cell wall properties, i.e., lignin content, microfibril angle, fiber length, lumen diameter, cell wall thickness, latewood proportion, and ring width, was available. A multiple linear regression analysis of all fiber and cell wall properties showed a significant, but not very strong effect on the variability of shear strength (R2 = 0.21). It is thus demonstrated that micro-structural variability plays a minor role in the variability of shear properties. By contrast, a multiple linear regression involving shear modulus, density, and slope of grain as three independent variables revealed an excellent possibility to model the variability of shear strength (R2 = 0.72). This study demonstrates the potential for non-destructive evaluation of the shear strength of solid wood.