Sulfur K-edge X-ray absorption near-edge structure (XANES) spectroscopy is used to measure how the speciation of sulfur compounds evolves within a warm-mix asphalt as a consequence of the Rolling Thin-Film Oven (RTFO) and Pressure Aging Vessel (PAV) oxidative aging procedures. Identifying the types of sulfur compounds present is important for quantifying the growth in polar sulfur-containing species that can alter the asphalt's mechanical properties over time. Elemental analysis indicates that the sulfur content of the asphalt holds constant at 5% during aging. XANES analysis indicates that thiophenic sulfur compounds are most prevalent (62%), followed by sulfide and elemental sulfur compounds. RTFO and PAV aging cause smaller and larger shifts from sulfide to sulfoxide. The amount of unreacted sulfide remains larger than the amount of sulfoxide, even with PAV aging. The XANES spectra lack features that would be expected if engine oil additives indicative of recycled engine oil bottoms were present. The results indicate the importance of including thiophene, sulfide, and sulfoxide chemistries within molecular asphalt models.

Q. Spring, "High-power light-emitting diode array design and assembly for practical photodynamic therapy research," J. AbstractSignificance: Commercial lasers, lamps, and light-emitting diode (LED) light sources have stimulated the clinical translation of photodynamic therapy (PDT). Yet, the continued exploration of new photosensitizers (PSs) for PDT often requires separate activation wavelengths for each agent being investigated. Customized light sources for such research frequently come at significant financial or technical cost, especially when compounded over many agents and wavelengths.Aim: LEDs offer potential as a cost-effective tool for new PS and multi-PS photodynamic research. A low-cost-per-wavelength tool leveraging high-power LEDs to facilitate efficient and versatile research is needed to further accelerate research in the field.Approach: We developed and validated a high-power LED array system for benchtop PDT with a modular design for efficient switching between wavelengths that overcome many challenges in light source design. We describe the assembly of a low-cost LED module plus the supporting infrastructure, software, and protocols to streamline typical in vitro PDT experimentation.Results: The LED array system is stable at intensities in excess of 100 mW∕cm 2 with 2.3% variation across the illumination field, competitive with other custom and commercial devices. To demonstrate efficacy and versatility, a primary ovarian cancer cell line was treated with two widely used PSs, aminolevulinic acid and verteporfin, using the LED modules at a clinically relevant 50 J∕cm 2 light dose that induced over 90% cell death for each treatment. Conclusions:Our work provides the community with a tool for new PS and multi-PS benchtop photodynamic research that, unlike most commercial light sources, affords the user a low barrier to entry and low-cost-per-wavelength with the goal of illuminating new insights at the forefront of PDT.

Photodynamic therapy (PDT) is an established therapeutic modality that uses nonionizing near‐infrared light to activate photocytotoxicity of endogenous or exogenous photosensitizers (PSs). An ongoing avenue of cancer research involves leveraging PDT to stimulate antitumor immune responses; however, these effects appear to be best elicited in low‐dose regimens that do not provide significant tumor reduction using conventional, nonspecific PSs. The loss of immune enhancement at higher PDT doses may arise in part from indiscriminate damage to local immune cell populations, including tumor‐infiltrating T cells. We previously introduced “tumor‐targeted, activatable photoimmunotherapy” (taPIT) using molecular‐targeted and cell‐activatable antibody–PS conjugates to realize precision tumor photodamage with microscale fidelity. Here, we investigate the immune cell sparing effect provided by taPIT in a 3D model of the tumor immune microenvironment. We report that high‐dose taPIT spares 25% of the local immune cell population, five times more than the conventional PDT regimen, in a 3D coculture model incorporating epithelial ovarian cancer cells and T cells. These findings suggest that the enhanced selectivity of taPIT may be utilized to achieve local tumor reduction with sparing of intratumor effector immune cells that would otherwise be lost if treated with conventional PDT.

Photosensitizer (PS)–antibody conjugates (photoimmunoconjugates, PICs) enable cancer cell‐targeted photodynamic therapy (PDT). Nonspecific chemical bioconjugation is widely used to synthesize PICs but gives rise to several shortcomings. The conjugates are heterogeneous, and the process is not easily reproducible. Moreover, modifications at or near the binding sites alter both binding affinity and specificity. To overcome these limitations, we introduce convergent assembly of PICs via a chemo‐enzymatic site‐specific approach. First, an antibody is conjugated to a clickable handle via site‐specific modification of glutamine (Gln) residues catalyzed by transglutaminase (TGase, EC 2.3.2.13). Second, the modified antibody intermediate is conjugated to a compatible chromophore via click chemistry. Utilizing cetuximab, we compared this site‐specific conjugation protocol to the nonspecific chemical acylation of amines using N‐hydroxysuccinimide (NHS) chemistry. Both the heavy and light chains were modified via the chemical route, whereas, only a glutamine 295 in the heavy chain was modified via chemo‐enzymatic conjugation. Furthermore, a 2.3‐fold increase in the number of bound antibodies per cell was observed for the site‐specific compared with nonspecific method, suggesting that multiple stochastic sites of modification perturb the antibody–antigen binding. Altogether, site‐specific bioconjugation leads to homogenous, reproducible and well‐defined PICs, conferring higher binding efficiency and probability of clinical success.

Photodynamic therapy (PDT) research would benefit from an automated, low-cost, and easy-to-use cell culture light treatment setup capable of illuminating multiple well replicates within standard multiwell plate formats. We present an LED-array suitable for performing high-throughput cell culture PDT experiments. The setup features a water-cooling loop to keep the LED-array temperature nearly constant, thus stabilizing the output power and spectrum. The setup also features two custom-made actuator arms, in combination with a pulse-width-modulation (PWM) technique, to achieve programmable and automatic light exposures for PDT. The setup operates at ~ 690 nm (676–702 nm, spectral output full-width half-maximum) and the array module can be readily adapted to other LED wavelengths. This system provides an illumination field with adjustable irradiance up to 400 mW/cm2 with relatively high spectral and power stability comparing with previously reported LED-based setups. The light doses provided by the LED array were validated with comparison to traditional laser PDT. This open-source illumination platform (including the detailed technical description, fabrication protocols, and parts list provided here) helps to make custom light sources more accessible and of practical use for photomedicine research.