High-throughput feedback-enabled optogenetic stimulation and spectroscopy in microwell plates

Abstract

The ability to perform sophisticated, high-throughput optogenetic experiments has been greatly enhanced by recent open-source illumination devices that allow independent programming of light patterns in single wells of microwell plates. However, there is currently a lack of instrumentation to monitor such experiments in real time, necessitating repeated transfers of the samples to stand-alone instruments and limiting the types of experiments that could be performed. Here we address this gap with the development of the optoPlateReader (oPR), an open-source, solid-state, compact device that allows automated optogenetic stimulation and spectroscopy in each well of a 96-well plate. The oPR integrates an optoPlate optical stimulation module with a module called the optoReader, an array of 96 photodiodes and LEDs that allows 96 parallel light measurements. The oPR was optimized for stimulation with blue light and for measurements of optical density and fluorescence. After calibration of all device components, we used the oPR to measure growth and to induce and measure fluorescent protein expression in E. coli. We further demonstrated how the optical read/write capabilities of the oPR permit computer-in-the-loop feedback control, where the current state of the sample can be used to adjust the optical stimulation parameters of the sample according to pre-defined feedback algorithms. The oPR will thus help realize an untapped potential for optogenetic experiments by enabling automated reading, writing, and feedback in microwell plates through open-source hardware that is accessible, customizable, and inexpensive.

Light-regulated gene expression in Bacteria: Fundamentals, advances, and perspectives

“…These approaches are particularly effective for photochromic receptors as their activity state can be bidirectionally changed by different light colors, as demonstrated for the widely used CcaRS TCS (Davidson et al, 2013;Milias-Argeitis et al, 2016;Chait et al, 2017;Steel et al, 2020). However, in principle other optogenetic circuits may also be controlled in feedback manner, as recently shown for pDusk, pDawn (Datta et al, 2022), and lightresponsive T7RNAP (Gutiérrez Mena et al, 2022).…”

Multiplexed dynamic control of temperature to probe and observe mammalian cells

“…Although designed for control of 96-well plates, the thermoPlate could be readily adapted to larger well formats where multiple reader/heater pairs would regulate a single well. Because the thermoPlate occupies only the top side of a well plate, it is also compatible with other instruments that stimulate or measure samples from the bottom of the plate [32][33][34] , opening the door for integration of additional inputs and outputs for thermoPlate experiments.…”

Lustro: High-Throughput Optogenetic Experiments Enabled by Automation and a Yeast Optogenetic Toolkit

“…However, these approaches still lack a method for high-throughput and rapid measurement of the optogenetic system response. The recent optoPlateReader 28 partially solves this problem, but requires the use of many biological replicates to obtain reliable data and lacks access to liquid handling capabilities, important for performing certain assays or long-term experiments.…”