We calculated the real and imaginary parts of the effective refractive index n eff of blood as functions of wavelength from 400 to 800 nm; we employed van de Hulst's theory, together with the anomalous diffraction approximation, for the calculation. We modelled blood as a mixture of plasma and erythrocytes. Our results indicate that erythrocyte orientation has a strong effect on n eff , making blood an optically anisotropic medium except when the erythrocytes are randomly oriented. In the case in which their symmetry axis is perpendicular to the wave vector, n eff equals the refractive index of plasma at certain wavelengths. Furthermore, the erythrocytes' shape affects their contribution to n eff in an important way, implying that studies on the effective refractive index of blood should avoid approximating them as spheres or spheroids. Finally, the effective refractive index of blood predicted by van de Hulst's theory is different from what would be obtained by averaging the refractive indices of its constituents weighted by volume; such a volume-weighted average is appropriate only for haemolysed blood. We then measured the real part of the refractive index of various blood solutions using two different experimental setups. One of the most important results of our expriment is that n eff is measurable to a good degree of precision even for undiluted blood, although not all measuring apparatuses are appropriate. The experimental data is self-consistent and in reasonable agreement with our theoretical calculations.

Lipid phase separation in cellular membranes is thought to play an important role in many biological functions. This has prompted the development of synthetic membranes to study lipid–lipid interactions in vitro, alongside optical microscopy techniques aimed at directly visualizing phase partitioning. In this context, there is a need to overcome the limitations of fluorescence microscopy, where added fluorophores can significantly perturb lipid packing. Raman-based optical imaging is a promising analytical tool for label-free chemically specific microscopy of lipid bilayers. In this work, we demonstrate the application of hyperspectral coherent Raman scattering microscopy combined with a quantitative unsupervised data analysis methodology developed in-house to visualize lipid partitioning in single planar membrane bilayers exhibiting liquid-ordered and liquid-disordered domains. Two home-built instruments were utilized, featuring coherent anti-Stokes Raman scattering and stimulated Raman scattering modalities. Ternary mixtures of dioleoylphosphatidylcholine, sphingomyelin, and cholesterol were used to form phase-separated domains. We show that domains are consistently resolved, both chemically and spatially, in a completely label-free manner. Quantitative Raman susceptibility spectra of the domains are provided alongside their spatially resolved concentration maps.

Light's internal reflectivity near a critical angle is very sensitive to the angle of incidence and the optical properties of the external medium near the interface. Novel applications in biology and medicine of subcritical internal reflection are being pursued. In many practical situations the refractive index of the external medium may vary with respect to its bulk value due to different physical phenomena at surfaces. Thus, there is a pressing need to understand the effects of a refractiveindex gradient at a surface for near-critical-angle reflection. In this work we investigate theoretically the reflectivity near the critical angle at an interface with glass assuming the external medium has a continuous depthdependent refractive index. We present graphs of the internal reflectivity as a function of the angle of incidence, which exhibit the effects of a refractive-index gradient at the interface. We analyse the behaviour of the reflectivity curves before total internal reflection is achieved. Our results provide insight into how one can recognise the existence of a refractive-index gradient at the interface and shed light on the viability of characterising it.

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