Protocol detail

Functional Imaging of Retinal Ganglion Cells

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For functional imaging experiments, a blue LED was used to stimulate the RGCs (Fig. 2a). The light from the LED was purified by a bandpass filter (FF02-447/60, Semrock) and directed to the mouse eye at a power level of ~60 μW/mm². The duration and timing of the blue light flashes were controlled by AO-TPEFM software. In the recording of calcium transients, a short pulse (~10 ms) of blue light was flashed at the mouse eye at 10 s intervals^{33 (link)}, while the two-photon fluorescence images were acquired at a frame rate of 3.8 Hz.

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Qin Z., He S., Yang C., Yung J.S., Chen C., Leung C.K., Liu K, & Qu J.Y. (2020). Adaptive optics two-photon microscopy enables near-diffraction-limited and functional retinal imaging in vivo. Light, Science & Applications, 9, 79.

Publication 2020

FF02-447/60

Calcium Fluorescence Frame Light Mouse Pulse Transients

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Variable analysis

independent variables

Duration and timing of the blue light flashes

dependent variables

Calcium transients

control variables

Power level of the blue light (~60 μW/mm^2)
Bandpass filter (FF02-447/60, Semrock) to purify the light from the LED
Frame rate of the two-photon fluorescence images (3.8 Hz)
Interval between the blue light flashes (10 s)

controls

Positive control: Not mentioned
Negative control: Not mentioned

Annotations

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