## 4 Discussion
These findings indicate that plaques in retardation images overall express a higher contrast than plaques visualized in intensity data when compared to surrounding neural tissue. The plaque sizes as measured by histology and the two different OCT contrasts differed from each other. The diameters of the plaques segmented in the retardation images showed a trend to be bigger than those segmented in the intensity data (Figure 5a). Plaques segmented in histology data, the gold standard, had a smaller plaque diameter when compared to plaques segmented in retardation and intensity images (Figure 5a versus Figure 7b). The minimum plaque size detected in histology was around 3 \(\upmu\)m, whereas the smallest plaques detected in OCT were around 12 \(\upmu\)m. While the detected size of plaques in all modalities did agree with earlier literature reports [44; 45; 46], the direct comparison between histology and OCT (Figures 5a and 7) showed that OCT seems to severely overestimate plaque size. Potential reasons for this could be the expansion (convolution) of imaged objects by the imaging system's point spread function (PSF), the inherently different contrast mechanisms and tissue shrinkage due to dehydration and fixation processes. Previous studies reported tissue shrinkage due to histologic workup up to 17% [47; 48]. Most importantly, the image resolution of OCT is rather poor when compared to histology. The observed size of a point-like object is described by the PSF of the imaging system. In an image, any object will be smeared out (convolved) with the PSF in x, y and z directions. The PSF in OCT is described by the coherence length in z-direction (which is determined by the covered spectral bandwidth in wavenumber space) and by the beam spot size in x-y direction [49]. Moreover, the beam spot size depends on the distance to the focal plane. Due to the limited resolution, multiple single plaques in close vicinity detected by histology might also be mistaken for a single, large plaque in OCT. In contrast, immunostaining visualizes amyloid-beta deposits of (almost) all shapes and sizes very specifically, and is expected to pick up much fainter signals and structures. OCT seems to visualize much fewer plaques (Figure 7a). Several methodological modifications may improve the performance of OCT for visualizing amyloid-beta plaques. In order to enhance the axial image resolution, a light source providing a broader bandwidth in wavenumber space (e.g., with a shorter central wavelength and/or broader wavelength coverage than the system used here) could be employed [32; 33]. Furthermore, objective lenses with higher magnification could be used for improving the lateral resolution. Note, however, that the use of objectives with a shorter focal length usually comes at the cost of a reduction of the FOV. As a logical next step, tissues of patients with different stages of the disease could be investigated to evaluate the time point at which plaques can be visualized with OCT. Further elucidation in the three-dimensional shape could be obtained by reconstructed histological serial sections similar to that shown in Figure 6a. As an advantage over histology, OCT can investigate ex-vivo cerebral tissue in a tissue-preserving manner, without any fixation, sectioning or staining. However, at the same time, OCT relies on intrinsic contrast and therefore does not require additional tissue processing steps. Hence, for some applications, multimodal OCT imaging might be a promising high throughput approach for the investigation of pathological alterations of the human brain.