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The hemaPEN is a liquid microsampling device for the reproducible collection and storage of blood samples as dried blood spots, for subsequent quantitative analysis. We examined the device’s ability to collect accurate and precise blood volumes, at different hematocrit levels, via in vitro studies using acetaminophen in human blood. We also investigated the impact of different user training approaches on device performance. The hemaPEN demonstrated acceptable volumetric accuracy and precision, regardless of the training medium used. Issues with apparent hematocrit-dependent bias were found to be associated with the extraction process, rather than the volumetric performance of the device. The hemaPEN is capable of readily producing high quality blood microsamples for reproducible and accurate quantitative bioanalysis.