We made several changes to earlier behavioral assay
protocols to improve the efficiency of the trials. Most significantly, we dramatically improved the scorpion image by
removing interference from the Plexiglas stage and glare from
direct IR projection of the recording camera. By using diffuse
IR from the side, the image cleared to a point that we could
easily detect scorpions using the public domain ImageJ image-
processing program. Once resolved from the background, the
animals were accurately tracked via Fiji’s Mtrack2 plugin.
Automated tracking greatly reduces scoring time and removes
the potential for human bias. We think similar tracking will be
useful for additional scorpion studies, including those
behaviorally testing for and identifying chemicals that make
up scorpion pheromone secretions (Taylor et al. 2012).