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Background subtraction was performed using the mean optical density value from 4 arbitrarily selected, vessel-free, areas (60×60 pixels).
Cut-off values were determined for each plate by using the mean optical density of all negative controls plus three standard deviations.
Cut-offs were calculated using the mean optical density readings from negative sample plus twice the standard deviation of negative samples.
In the same graph, we show that comparable results can be obtained for the sampling volume using the mean optical properties of excitation and Stokes shifted wavelength in the simulation while neglecting the Stokes shift.
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GAP-43 protein expression in the sciatic nerve was also measured and processed by this method using the average optical density (mean density).
The mean optical density (OD) was used for quantification using a routine microtitre plate reader at a wavelength of 570 nm.
Second, data from the air mass 1.5 spectrum are used as weight factors to calculate the mean optical transmittance.
The mean optical density from the control slides was used to obtain a baseline of endogenous fluorescence.
Concentrations were calculated using a standard curve generated for each ELISA plate in duplicates and fitted with linear regression of the expected concentrations against the mean optical density.
Graphed data are presented as the mean optical density (group means mGluR5/actin immunolevels) ± the standard error of the mean (SEM).
The mean optical density (490 nm) or the mean relative light units (RLU) obtained with the PPD+/PPD− healthy individuals + 3 standard deviations (SD) was used as cut-off to determine positive reactivity.
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