The sensing performance of AlGaN/GaN MOS-HEMT biosensors for neutral biomolecules was investigated by Silvaco TCAD. In order to achieve excellent sensing sensitivity, the gate length (1–4 µm) and AlGaN barrier layer thickness (10–30 nm) of the MOS-HEMT were optimized. The results show that by keeping the sensing area constant, the sensing performance of MOS-HEMT decreases with the increase of the gate length. However, the sensing sensitivity increases with the increase of gate length by keeping the gate dielectric length at 0.5 µm. Moreover, the trend of ∆IDS change slows down as the gate length increases. In addition, the relatively thinner AlGaN barrier layer can optimize the sensing performance of the MOS-HEMT biosensor. Ultimately, the optimal sensitivity was obtained of MOS-HEMT with a 1.5-µm-long gate and 10-nm-thick AlGaN barrier layer. For casein biomolecules, the ∆IDS of MOS-HEMT is 196.8 mA/mm. This study offers crucial validation for the advancement of GaN-based MOS-HEMT biosensors, significantly contributing to the field’s progress.