Near-Field IR Orientational Spectroscopy of Silk

Orientational dependence of the IR absorbing amide bands of silk is demonstrated from two orthogonal longitudinal and transverse microtome slices only $\sim 100$~nm thick. A scanning near-field optical microscopy (SNOM) which preferentially probes orientation perpendicular to the sample's surface was used. Spatial resolution of silk-epoxy boundary was defined with a $\sim 100$~nm resolution while the spectra were collected by a $\sim 10$~nm tip. Ratio of the absorbance of the amide-II C-N at 1512~cm$^{-1}$ and amide-I C=O $\beta$-sheets at 1628~cm$^{-1}$ 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 slice cut orientation, which is equivalent to the four polarisation angles absorbance measurement is proposed.