Figure 1

IR-LEGO experimental setup combined with confocal spinning disk for live imaging

(A) IR-LEGO system schematic. The infrared photo-irradiation device (IR-LEGO module in light red) is coupled to a spinning disk microscope dedicated to fluorescence live imaging (spinning disk confocal module in gray). IR-LEGO module consists in an infrared laser diode, a galvanometric head mirror to align the laser in the center of the field, a mirror, a dichroic mirror D1 and an IR compatible objective lens, to focalize the laser on the sample. Spinning disk confocal module contains an inverted microscope equipped with an imaging laser combiner composed of 4 diode lasers, a Nipkow spinning head (Yokogawa CSU X1), a fast SCMOS camera and a galvanometric stage for fast image acquisition.

(B) A schematic of the sample setup for IR-LEGO using a 40 X APO IR water objective. The magnification shows the laser target in the middle of the zebrafish embryo telencephalon required here for the experiments.

(C) IR irradiation of "upconverted phosphor particles" imaged in transmission. "Upconverted phosphor particles" were irradiated with a power of 90% IR laser, inducing an emission wavelength of 545 nm. The irradiation diameter on the calibration sample is estimated to 34 μm (black outlined circle, grey line shows the diameter).

(D and E) Experimental system for synchronization of IR irradiation (D) and live imaging (E) conditions. Synchronization timings of all laser-induced heating components followed by (E) imaging chromatogram for each component to reduce phototoxicity. These timings are driven by Metamorph ("synchronization card").