# Can OCT Angiography Be Made a Quantitative Blood Measurement Tool?
## 2 OCTA Fundamentals
### Light Scattering from Red Blood Cells  
What are the physical properties of RBCs that enable their detection by OCTA? RBC scattering and absorption properties derive from the presence of hemoglobin and its complex refractive index [26]. Major absorption bands of hemoglobin, related to the imaginary part of the complex refractive index, predominate at visible and shorter wavelengths, while hemoglobin absorption becomes negligible at near-infrared wavelengths, where scattering dominates. The light scattering properties of individual RBCs are determined by the refractive index contrast with respect to the surrounding plasma, as well as their shape and size relative to the medium wavelength. The real part of the complex refractive index, or refractive index, of hemoglobin is larger by \(\sim\)3-6% relative to the surrounding plasma [26; 27; 28]. RBCs are biconcave disks (Figure 2), with a diameter of 6-8 \(\upmu\)m and thickness of \(\sim\)2 \(\upmu\)m, although their shape changes under external stress. Due to the large volume fraction of RBCs and their refractive index mismatch relative to plasma, RBCs are the main scattering constituent in blood [27; 29; 30]. The scattering properties of both individual RBCs and ensembles of RBCs are important in OCTA. Due to their irregular shape, the probability of light scattering in a given direction for a particular RBC depends on both its orientation and the direction of incident light. An ensemble of RBCs with different orientations can be characterized by a scattering coefficient (\(\mu_{\text{s}}\)), the scattering probability per unit distance; a scattering phase function (\(P(\theta)\)), the probability of scattering in a given elevation direction \(\theta\) per unit solid angle; and a scattering anisotropy (\(g=E[\cos(\theta)]\)), the expectation or average (\(E[\,]\)) of \(\cos(\theta)\) over solid angle. These parameters characterize scattering of whole blood, which comprises an ensemble volume of RBCs with random orientations. In whole blood, empirically measured g and \(\mu_{\text{s}}\) include dependent and multiple scattering effects [31]. With a hematocrit of around 45%, whole blood is found to be highly forward scattering between 750 and 950 nm, with a scattering coefficient  
Figure 2: Single and multiple scattering in the OCTA of capillaries versus macrovascularture. (**A**) Blood has a high scattering anisotropy, leading to a high probability of detecting multiple scattered light paths; (**B**) For capillaries, dynamic RBC (red blood cell) forward scattering precedes or follows static tissue backscattering, which leads to ”multiple scattering” tails; (**C**) In large vessels, the backscattering cross-section is determined by the shear-induced orientation of RBCs with their flat face parallel to the shear force. If the vessel lumen exceeds a scattering length, multiple intravascular dynamic scattering events (green) before detection are likely; (**D**) Cross-sectional OCT (Optical Coherence Tomography) angiogram of the mouse brain at 1300 nm (complex interframe subtraction method) with a qualitative colorbar showing the balance of backscattered light (blue) and multiple scattered light (green) in a capillary (left) and macrovessel (right). (\(\mu_{s}\)) between 65 and 80 mm\({}^{-1}\), and anisotropy (g) between 0.97 and 0.99 [29; 32; 33; 34]. Exemplary phase functions [30; 32; 35] for tissue (Henyey-Greenstein with \(g~{}=~{}0.9\)) and blood (Gegenbauer-Kernel with \(g~{}=~{}0.972\) and \(\alpha~{}=~{}0.49\)[36]) are shown in Figure 2A on a logarithmic scale. Tissue has a higher probability of back scattering than blood, while blood is considerably more forward scattering. In OCTA (Figure 2B,C), detected light ideally results from paths with single RBC backscattering (\(\theta=180^{\circ}\)) events (blue). However, the high RBC anisotropy (Figure 2A) makes detection of multiple scattered light (green) likely. Probable light paths can be understood through the principles of radiative transport.