Complete tumor removal during breast-conserving surgery remains challenging due to the lack of optimal intraoperative margin assessment techniques. Here, we use hyperspectral imaging for tumor detection in fresh breast tissue. We evaluated different wavelength ranges and two classification algorithms; a pixel-wise classification algorithm and a convolutional neural network that combines spectral and spatial information. The highest classification performance was obtained using the full wavelength range (450-1650 nm). Adding spatial information mainly improved the differentiation of tissue classes within the malignant and healthy classes. High sensitivity and specificity were accomplished, which offers potential for hyperspectral imaging as a margin assessment technique to improve surgical outcome.

Light fractionation with dark periods of the order of hours has been shown to considerably increase the efficacy of 5‐aminolevulinic acid‐photodynamic therapy (ALA‐PDT). Recent investigations have suggested that this increase may be due to the resynthesis of protoporphyrin IX (PpIX) during the dark period following the first illumination that is then utilized in the second light fraction. We have investigated the kinetics of PpIX fluorescence and PDT‐induced damage during PDT in the normal skin of the SKH1 HR hairless mouse. A single illumination (514 nm), with light fluences of 5, 10 and 50 J cm−2 was performed 4 h after the application of 20% ALA, to determine the effect of PDT on the synthesis of PpIX. Results show that the kinetics of PpIX fluorescence after illumination are dependent on the fluence delivered; the resynthesis of PpIX is progressively inhibited following fluences above 10 J cm−2. In order to determine the influence of the PpIX fluorescence intensity at the time of the second illumination on the visual skin damage, 5 + 95 and 50 + 50 J cm−2 (when significantly less PpIX fluorescence is present before the second illumination), were delivered with a dark interval of 2 h between light fractions. Each scheme was compared to illumination with 100 J cm−2 in a single fraction delivered 4 or 6 h after the application of ALA. As we have shown previously greater skin damage results when an equal light fluence is delivered in two fractions. However, significantly more damage results when 5 J cm−2 is delivered in the first light fraction. Also, delivering 5 J cm−2 at 5 mW cm−2+ 95 J cm−2 at 50 mW cm−2 results in a reduction in visual skin damage from that obtained with 5 + 95 J cm−2 at 50 mW cm−2. A similar reduction in damage is observed if 5 + 45 J cm−2 are delivered at 50 mW cm−2. PpIX photoproducts are formed during illumination and subsequently photobleached. PpIX photoproducts do not dissipate in the 2 h dark interval between illuminations.

DPS is a new optical technique allowing the noninvasive study of endobronchial tumor hypoxia.

A fractionated illumination scheme in which a cumulative fluence of 100 J cm 22 is delivered in two equal light fractions separated by a dark interval of 2 h has been shown to considerably increase the efficacy of 5-aminolevulinic acid (ALA)-photodynamic therapy (PDT). The efficacy of such a scheme is further increased if the fluence of the first light fraction is reduced to 5 J cm 22 . We have investigated the relationship between the PDT response and the kinetics of protoporphyrin IX (PpIX) fluorescence in the SKH1 HR hairless mouse for first fraction fluences below 5 J cm 22 delivered 4 h after the application of ALA and 10 J cm 22 delivered 2 h after the application of ALA. Illumination is performed using 514 nm at a fluence rate of 50 mW cm 22 . Reducing the fluence of the first fraction to 2.5 J cm 22 does not result in significantly different visual skin damage. The PDT response, however, is significantly reduced if the fluence is lowered to 1 J cm 22 , but this illumination scheme (1 1 99 J cm 22 ) remains significantly more effective than a single illumination of 100 J cm 22 . A first light fraction of 10 J cm 22 can be delivered 2 h earlier, 2 h after the application of ALA, without significant reduction in the PDT response compared with 5 1 95 J cm 22 delivered 4 and 6 h after the application of ALA. The kinetics of PpIX fluorescence are consistent with those reported previously by us and do not explain the significant increase in PDT response with a twofold illumination scheme. Histological sections of the illuminated volume showed a trend toward increasing extent and depth of necrosis for the two-fold illumination scheme in which the first light fraction is 5 J cm 22 , compared with a single illumination scheme.

We present a mathematical layer model to quantitatively calculate the diffusion of 5‐aminolevulinic acid (ALA) in the skin in vivo, its uptake into the cells and its conversion to protoporphyrin IX (PpIX) and subsequently to heme. The model is a modification and extension of a recently presented three‐compartment model. The diffusion of ALA in the skin (epidermis, dermis) is described by the time‐dependent diffusion equation, and the sink in this equation accounts for ALA uptake in the cells. As boundary conditions, we use the ALA flux across the human stratum corneum (SC) in vitro during passive or iontophoretic ALA delivery as measured in vitro. Besides the diffusion equation, the model includes three additional equations, similar in form to those of the three‐compartment model but with a different interpretation. Our additional equations are supposed to describe, respectively, the conversion of ALA in the cytoplasm to some intermediate compound in the mitochondria and the conversion of the latter to PpIX and of PpIX to heme. The first conversion is a process of the Michaelis–Menten type, the other two are first‐order rate processes. When fitted to the published data of PpIX fluorescence from normal human skin following iontophoresis of ALA, the model yields the tissue concentration of PpIX as a function of time after ALA application. The computed concentrations are in good agreement with the published phototoxic concentrations of PpIX in the tissues obtained from extraction. The model parameters obtained from the fit are subsequently used to compute the PpIX concentration in normal human skin after 4 h topical application of 10, 20 and 40% ALA. This again yields the PpIX concentrations in tissue, in good agreement with the published values. The saturation of the PpIX concentration as a function of applied ALA concentration is calculated and agrees with clinical observations on the effectiveness of photodynamic therapy. Photobleaching is simulated, with subsequent resynthesis of PpIX in qualitative agreement with experiment. Finally, the model predicts that only 2.5–3.5% of the ALA entering the skin after passing the SC is converted to PpIX. The layered model is a considerable simplification of real skin, but its successful qualitative and quantitative reproduction of experimental data may encourage further studies to test and refine the model to improve our understanding of the kinetics of ALA and the synthesis of PpIX in the skin.