Pm100usb

The optical fiber was connected using a pigtail rotary joint (Thorlabs) to a 470 nm laser (Ikecool, Anaheim, CA, USA) or a 589 nm laser (Laserglow, ON, Canada) driven by a Grass S88X that generated the stimulation trains (10 s train, 10 ms pulses, 20 Hz; Caggiano et al., 2018 (link); Josset et al., 2018 (link)). To visualize optogenetic stimulation, a small (diameter 0.5 cm), low-power (0.13 W) red LED that received a copy of the stimulation trains was placed in the field of view of the camera placed above the open field. The 470 nm light source was adjusted to 6–27% of laser power and the 589 nm to 40–53% of laser power. The corresponding power measured at the fiber tip with a power meter (PM100USB, Thorlabs) was 0.1–16.0 mW for the 470 nm laser and 1.7–9.4 mW for the 589 nm laser.

The curing lamp used in this study is X-Lite-II, which has a broad emission wavelength of 385–515 nm with a manufacturer specified intensity of 1700 mW/cm². The lamp head diameter was 8 mm and was placed on a linear translation stage. To measure the intensity profile of the lamp, a power meter based setup is used as shown in Fig. 1. The sensor used was S142C (Thorlabs) which has a measurement range of (300–700 nm) connected to a power energy meter PM100USB (Thorlabs). To measure the lamp profile, a pin-holed mount was placed on the power meter sensor (where pin hole is in the middle of the sensor) and was scanned edge to edge of the lamp head.Figure 1

Experimental setup to measure the intensity profile of the curing lamp.

The optical setup for writing the aligning direction on photoalignment layers comprises a UV light‐emitting diode (365 nm), an SLM (SM7‐405, Sicube Photonics Co., Ltd.), and a motorized rotational stage (K10CR1/M, Thorlabs) furnished with a UV polarizer (LPUV‐100‐MP2, Thorlabs). The SLM is a digital micromirror device that has a wide extended graphics array (WXGA, 1280 × 800 pixels). The polarizer works synchronously with the SLM. Target substrates were mounted on a three‐axis translation stage (DT12XYZ, Thorlabs) to align with the optical setup. A rail (RLA600/M, Thorlabs) and rail carriers (RC4, Thorlabs) were used to adjust the distance between substrates and the optical setup. The intensity of UV light at substrates was controlled to be 2.2 mW cm^–2. It was measured using a power meter (PM100USB, Thorlabs). The maximum dispersion of the light intensity across the center to the boundary was controlled to be below 7%.

All microscopy was performed with a Nikon Ti-E inverted microscope, operated using NIS-Elements. Light sources for bright field and fluorescence imaging were a halogen lamp and an LED (pE-2, CoolLED), respectively. 365 nm-light was for excitation in NAD(P)H-channel with a filter set comprising a 350/50-nm band-pass filter, a 409-nm beam-splitter and a 435/40-nm emission filter while 470 nm-light was for the fluorescence dye with a filter set comprising a 470/40-nm band-pass filter, a 495-nm beam-splitter and a 525/50-nm emission filter. A wavelength-optimized 100x objective (MRF02900, Nikon) and camera (DU-897 EX, Andor) were used for image acquisition. The EM Gain Multiplier of the camera was disabled and the Readout Mode was set to 1 MHz to reduce the camera readout noise. The baseline level of camera was set to 500 at −75 °C to avoid negative intensity value. Exposure energies were determined as exposure time (set in NIS elements) multiplied by exposure power, which was measured with a power meter (PM100USB, Thorlabs) and a power sensor (S120VC, Thorlabs), located at the specimen plane. The Nikon Perfect Focus System was used in all time-lapse movies to ensure focusing. Images in bright field channel were focused 1.05 μm higher than that in the fluorescence channels to facilitate cell segmentation.

The current pulse provided to LED sources was measured through the integrated output of the DLD, whereas the emitted optical pulses were characterized using a Si-based photodiode (DET36A, Thorlabs, Newton, NJ, USA; rise/fall time: 14 ns). All data were recorded by the digital oscilloscope of the PA system (NI USB-5133). The emission spectra of LEDs were determined by a spectrograph (getSpec 2048, getAmo, Sofia, Bulgaria) using 1 nm wavelength steps. The average power of the LEDs during the pulsed operation was measured using a sensitive thermal power sensor head (S405C, Thorlabs, Newton, NJ, USA) connected to a USB power meter interface (PM100USB, Thorlabs, Newton, NJ, USA). For each measurement, the sensor head was placed at the bottom of an empty sample container, with its front side facing directly the 10 mm diameter circular window. In every case, the LED was positioned at a fixed distance below the surface of the coverslip glass. The two LED sources operated one at a time, and were placed at identical positions to achieve similar illumination conditions on the phantoms. The optical power on the sample was controlled using a set of neutral density filters (NDK01, Thorlabs, Newton, NJ, USA). Absorption spectra of phantom samples were measured by a UV-VIS spectrophotometer (BK-D590, Biobase, Jinan, China) using 1 cm path plastic cuvettes and a wavelength step of 1 nm.

The steady-state singlet oxygen emission measurements were performed
on a previously reported custom-built setup utilizing a slightly modified
experimental procedure.^{59 (link)−61 (link)} Perinaphthenone was used as a
reference with reported φ_Δ(¹O₂) = 0.98 ± 0.07 in air-saturated ACN and φ_Δ(¹O₂) = 0.97 ± 0.04 in air-saturated
CD₃OD.^{48 (link),62 (link)} Ru(bpy)₃Cl₂ was used as a validation compound with reported φ_Δ(¹O₂) = 0.57 ± 0.06 in air-saturated ACN
and φ_Δ(¹O₂) = 0.73 ±
0.12 in air-saturated CD₃OD.^{49 (link),63 (link)} All the compounds
were dissolved in ACN or CD₃OD (V = 3
mL) and transferred into a macro fluorescence cuvette from Firefly
(lightpaths: 1 cm × 1 cm). The irradiation of samples was done
at 298 K using a 450 nm LRD-0450 Laserglow fiber-coupled laser set
to 80 mW at the cuvette with help of a PM100USB Thorlabs power meter.
The UV–vis and NIR spectra were recorded at 298 K with Agilent
Cary 60 UV–vis and Avantes NIR256–1.7TEC spectrometers,
respectively. The NIR emission spectra were acquired within 20 s.
All the spectral data were processed with OriginPro 9.1 and Microsoft
Office Excel 2016.