The Orbitrap mass analyzer can provide high mass accuracy and throughput, which has significantly improved proteome research and made this type of instrumentation one of the most frequently applied in proteomics. The efficient use of Orbitrap mass spectrometers requires training. Students in the field of proteomics can benefit from a deeper understanding of the Orbitrap technology to comprehend mass spectral interpretation, troubleshooting, and judgment of experimental settings. Unfortunately, the cost of high-end mass spectrometers limits the implementation of this type of equipment in educational laboratories. Guided by these concerns, we developed an eLearning web application called HUMOS aimed to help teach Orbitrap mass spectrometry. Although a typical proteomics experiment includes the use of several different technologies, such as liquid chromatography, mass spectrometry, and bioinformatics, the learning objectives of HUMOS are focused on mass spectrometry. HUMOS models a mass spectrum of a peptide mixture, allowing us to investigate the influence of mass spectral acquisition parameters. By changing the parameters and observing the differences, students can learn more about the mass spectral resolution, duty cycle, throughput of the analysis, ion accumulation, and spectral dynamic range and get familiar with advanced spectral acquisition methods, such as BoxCar. HUMOS is an open-source software published under the Apache license; the live installation is available at http://humos.bmb.sdu.dk.