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. 2020 Oct;69(4):313-316.
doi: 10.2144/btn-2020-0077. Epub 2020 Jul 29.

Automated calibration of optoPlate LEDs to reduce light dose variation in optogenetic experiments

Edvard Os Grødem  1 , Kieran Sweeney  1 , Megan N McClean  1   2
Affiliations

Automated calibration of optoPlate LEDs to reduce light dose variation in optogenetic experiments

Edvard Os Grødem et al. Biotechniques. 2020 Oct.

Abstract

Optogenetic systems use light to precisely control and investigate cellular processes. Until recently, there had been few instruments available for applying controlled light doses to cultures of cells. The optoPlate, a programmable array of 192 LEDs, was developed to meet this need. However, LED performance varies, and without calibration there are substantial brightness differences between LEDs on an optoPlate. Here we present a method for calibrating an optoPlate that uses a programmable microscope stage and optical power meter to automatically measure all 192 LEDs of an optoPlate. The resulting brightness measurements are used to calculate calibration values that tune the electrical current supplied to each optoPlate LED to reduce brightness variation in optogenetic experiments.

Keywords: calibration; optoPlate; optogenetics.

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Conflict of interest statement

Financial & competing interests disclosure

This work was supported by the National Institute of Allergy and Infectious Diseases (R21AI135166) and the National Institute of General Medical Sciences (R35GM128873). K Sweeney is supported by a Genomic Sciences Training Program NHGRI Training Grant (5T32HG002760). MN McClean holds a Career Award at the Scientific Interface from the Burroughs Wellcome Fund. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Figures

Figure 1.
Figure 1.. Measuring optoPlate LEDs.
(A) Our optoPlate calibration method involves a measurement process in which an optical power meter is rastered across the optoPlate over a series of four passes in which one LED is active in alternating wells. (B) This generates irradiance measurements from which it is easy to identify each LED and measure its brightness.
Figure 2.
Figure 2.. Calibration reduces optoPlate LED irradiance variation.
(A) Measurements of LED irradiances before and after calibration show that calibration greatly reduces LED brightness variation. (B) A histogram depicting the same data shows that calibration reduces LED variation by dimming the brightest LEDs to match the irradiance of the dimmest LEDs. (C) Experimental light doses can vary strongly for an uncalibrated optoPlate; these differences almost disappear following calibration.
Figure 3.
Figure 3.. Calibration validation.
(A) The dose–response relationship between input value and light dose for a calibrated optoPlate. Four measurements were made per input value. Standard deviation of the measured irradiances is small and obscured by the markers. (B) Brightness variation for an optoPlate at a fixed brightness level before and after calibration. All optoPlate wells were measured for each condition. Asterisks indicate p < 0.001 as calculated by Levene's test for equality of variances.
See this image and copyright information in PMC

References

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