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A cohort of 100 bees was marked with blue paint on the thorax at emergence, caged with honey ad libitum as a food source, and kept in an incubator in the dark for the duration of the experiment.
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Bees were marked with individually numbered tags within 1 day of emergence from pupae.
50 bees were marked with pink paint and had both antennae amputated with a sterile blade; 50 bees were marked with yellow paint and had the right antenna amputated; 50 bees were marked with green paint, and had the left antenna amputated; the last 50 bees were marked with blue paint and had both antenna intact, serving as control bees.
Parallel to this, another 100 bees were marked with white paint at emergence and returned to the hive, representing the control bee cohort (hive bees).
These bees were marked with enamel paint, released, and not counted in subsequent trials to avoid pseudoreplication.
Worker bees were marked with paint on their thorax when just emerging from the cells and then placed back into colonies.
Treated bees were marked with a dot of Testor's paint on their thorax and maintained individually in 10x10x7 cm Plexigas cages in an incubator under red light at 33°C, 50%RH for 6 hours, with 50% sucrose.
Each individual bee was marked with a unique tag.
Workers from a commercially obtained Bombus terrestris colony (Syngenta Bioline Bees, Weert, The Netherlands) were marked with individually numbered tags within 1 day of emergence.
For the second paradigm (associative scent learning), another 100 bees were marked at emergence with paint and returned to the hive until collection.
For the behavioral assay, all bees were marked on their thorax with a dot of enamel paint, except for the bee used for the behavioral assay.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com