Orientational dependence of the IR absorbing amide bands of silk is demonstrated from two orthogonal longitudinal and transverse microtome slices with a thickness of only ∼100 nm. Scanning near-field optical microscopy (SNOM) which preferentially probes orientation perpendicular to the sample’s surface was used. Spatial resolution of the silk–epoxy boundary was ∼100 nm resolution, while the spectra were collected by a ∼10 nm tip. Ratio of the absorbance of the amide-II C-N at 1512 cm −1−1 and amide-I C=O ββ -sheets at 1628 cm −1−1 showed sensitivity of SNOM to the molecular orientation. SNOM characterisation is complimentary to the far-field absorbance which is sensitive to the in-plane polarisation. Volumes with cross sections smaller than 100 nm can be characterised for molecular orientation. A method of absorbance measurements at four angles of the slice cut orientation, which is equivalent to the four polarisation angles absorbance measurement, is proposed.