Paper-based analytical
devices (PADs) employing colorimetric detection
and smartphone images have gained wider acceptance in a variety of
measurement applications. PADs are primarily meant to be used in field
settings where assay and imaging conditions greatly vary, resulting
in less accurate results. Recently, machine-learning (ML)-assisted
models have been used in image analysis. We evaluated a combination
of four ML models—logistic regression, support vector machine
(SVM), random forest, and artificial neural network (ANN)—as
well as three image color spaces, RGB, HSV, and LAB, for their ability
to accurately predict analyte concentrations. We used images of PADs
taken at varying lighting conditions, with different cameras and users
for food color and enzyme inhibition assays to create training and
test datasets. The prediction accuracy was higher for food color than
enzyme inhibition assays in most of the ML models and color space
combinations. All models better predicted coarse-level classifications
than fine-grained concentration classes. ML models using the sample
color along with a reference color increased the models’ ability
to predict the result in which the reference color may have partially
factored out the variation in ambient assay and imaging conditions.
The best concentration class prediction accuracy obtained for food
color was 0.966 when using the ANN model and LAB color space. The
accuracy for enzyme inhibition assay was 0.908 when using the SVM
model and LAB color space. Appropriate models and color space combinations
can be useful to analyze large numbers of samples on PADs as a powerful
low-cost quick field-testing tool.