[Paper Review] Nanoscale optical and structural characterisation of silk

Meguya Ryu, Reo Honda+11 more|arXiv (Cornell University)|Feb 7, 2019

Silk-based biomaterials and applicationsMaterials Science3 citations

TL;DR

This study achieves sub-10 nm spatial resolution in hyperspectral infrared (IR) mapping of silk using nano-thin (100 nm) longitudinal microtome slices, enabling direct measurement of nanoscale optical anisotropy. It demonstrates reliable detection of only 1–2% transmission changes due to molecular orientation in amide bands, validating that structural and optical properties of silk remain intact at the nanoscale.

ABSTRACT

Background: Nanoscale composition of silk defining its unique properties via a hierarchical structural anisotropy has to be analysed at the highest spatial resolution of tens-of-nanometers corresponding to the size of fibrils made of b-sheets, which are the crystalline building blocks of silk. Results: Nanoscale optical and structural properties of silk have been measured from 100-nm thick longitudinal slices of silk fibers with ~10 nm resolution, the highest so far. Optical sub-wavelength resolution in hyperspectral mapping of absorbance and molecular orientation were carried out for comparison at IR wavelengths 2-10 micrometers using synchrotron radiation. Conclusion: Reliable distinction of transmission changes by only 1-2% due to anisotropy of amide bands was obtained from nano-thin slices of silk.

Motivation & Objective

• To achieve nanoscale spatial resolution in optical and structural characterization of silk, particularly at the level of β-sheet fibrils.
• To investigate whether sub-wavelength resolution can reliably detect anisotropic absorbance in silk due to molecular orientation of amide bands.
• To validate that ultra-thin (100 nm) microtome slices preserve the intrinsic optical and structural properties of soft biopolymers like silk.
• To compare far-field and near-field IR measurements for assessing absorbance and birefringence at the nanoscale.
• To determine if nano-slices eliminate optical averaging effects present in bulk or thicker samples, enabling direct measurement of intrinsic optical properties.

Proposed method

• Preparation of 100-nm-thick longitudinal cross-sections of silk fibers using an ultramicrotome to achieve nanoscale thickness for minimal optical averaging.
• Employment of synchrotron-based near-field scanning optical microscopy (SNOM) with a tip resolution of ~10 nm for hyperspectral IR mapping in the 2–10 µm range.
• Hyperspectral absorbance and retardance measurements were performed at IR wavelengths to assess molecular orientation anisotropy via dichroism Δ" = k(κ∥ − κ⊥)d.
• Comparison of absorbance spectra from SNOM (near-field) and far-field FTIR/ATR-FTIR techniques to validate consistency across measurement modalities.
• Use of the equation T∥/T⊥ = exp(−2Δ") to quantify transmission differences between parallel and perpendicular polarization states, linking dichroism to measurable transmission changes.
• Analysis of amide I, II, and A bands to assess orientation-dependent absorbance and birefringence in relation to structural anisotropy.

Experimental results

Research questions

• RQ1 Can sub-10 nm spatial resolution be achieved in hyperspectral IR mapping of silk to resolve nanoscale structural and optical anisotropy?
• RQ2 What is the magnitude of absorbance anisotropy (dichroism) in 100-nm-thick silk slices, and can it be reliably measured at the nanoscale?
• RQ3 Do ultra-thin microtome slices of silk preserve the intrinsic optical and structural properties of the bulk material?
• RQ4 How do near-field (SNOM) and far-field IR measurements compare in terms of spectral features and sensitivity to molecular orientation?
• RQ5 To what extent do transmission changes due to molecular anisotropy (e.g., T∥/T⊥) fall within the detectable range of current nanoscale IR techniques?

Key findings

• The study achieved a spatial resolution of ~10 nm using SNOM, the highest reported to date for nanoscale optical characterization of silk.
• Absorbance anisotropy due to molecular orientation was reliably measured, with only 1–2% transmission changes between parallel and perpendicular polarization states.
• For Amide I, Δ" ≃ 0.027, resulting in a transmission ratio T∥/T⊥ ≈ 97.4%, confirming measurable but small anisotropy.
• For Amide A, Δ" ≃ 0.014, leading to T∥/T⊥ ≈ 98.6%, indicating minimal but detectable dichroism.
• The far-field and near-field absorbance spectra were quantitatively comparable, validating the reliability of SNOM measurements on nano-slices.
• The structural and optical properties of silk, including birefringence (Δn ≈ 4×10⁻³) and absorbance, were preserved in 100-nm-thick microtome slices, confirming minimal artefacts from sample preparation.