Source Paper
Action Spectrum for Melatonin Regulation in Humans: Evidence for a Novel Circadian Photoreceptor
George C. Brainard, John P. Hanifin, et al.
Use this page as an execution guide, then fall back to the source paper whenever you need exact exclusions, dosing details, or assay-specific caveats.
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- Verify the animal model, intervention setup, and collection timepoints against the source paper.
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Protocol Steps
Start here. The step list is optimized for running the experiment, with direct vendor links available inline when you need to source a cited item.
Subject recruitment and screening
Recruit healthy subjects with normal color vision. Study population consisted of 37 females and 35 males with mean age of 24.5 ± 0.3 years
Note: Subjects must have normal color vision and be in good health
View evidence from paper
“Subjects (37 females, 35 males, mean age of 24.5 ± 0.3 years) were healthy and had normal color vision”
Pupil dilation
Dilate subjects' pupils prior to light exposure
Note: Pupils must be dilated during monochromatic light exposure
View evidence from paper
“Full-field, monochromatic light exposures took place between 2:00 and 3:30 A.M. while subjects' pupils were dilated”
Baseline blood sample collection
Collect blood sample before light exposure for baseline melatonin quantification
Note: Sample collected during nighttime hours
View evidence from paper
“Blood samples collected before and after light exposures were quantified for melatonin”
Monochromatic light exposure
Expose subject to full-field monochromatic light at controlled irradiance. Wavelengths tested ranged from 420 to 600 nm
Note: Each subject tested with at least seven different irradiances of one wavelength. Minimum 1 week between each nighttime exposure
View evidence from paper
“Full-field, monochromatic light exposures took place between 2:00 and 3:30 A.M. while subjects' pupils were dilated”
Post-exposure blood sample collection
Collect blood sample after light exposure for melatonin quantification
Note: Sample collected immediately after light exposure period
View evidence from paper
“Blood samples collected before and after light exposures were quantified for melatonin”
Melatonin quantification
Quantify melatonin levels in blood samples collected before and after light exposure
Note: Method of quantification not specified in text
View evidence from paper
“Blood samples collected before and after light exposures were quantified for melatonin”
Repeat testing with different wavelengths and irradiances
Conduct additional melatonin suppression tests across wavelength range of 420 to 600 nm with varying irradiances
Note: Total of 627 nighttime melatonin suppression tests completed across all subjects and wavelengths
View evidence from paper
“Each subject was tested with at least seven different irradiances of one wavelength with a minimum of 1 week between each nighttime exposure”
Data fitting and action spectrum construction
Fit melatonin suppression data to sigmoidal fluence-response curves and construct action spectrum by fitting data to opsin template
Note: Eight univariant sigmoidal curves generated with R² = 0.81–0.95; action spectrum fit opsin template with R² = 0.91
View evidence from paper
“The data were fit to eight univariant, sigmoidal fluence–response curves (R² = 0.81–0.95). The action spectrum constructed from these data fit an opsin template (R² = 0.91)”