Alzheimer's disease (AD) is a complex pathological process that is one of the leading causes of dementia globally. The demand for diagnostic tools that are minimally invasive, timely, and accurate is on the rise. Total tau (T-Tau) protein in blood serum is a promising biomarker for predicting early-stage AD diagnosis. In this study, the hexagonal boron nitride (HBN) based immunosensor platform was developed to detect T-Tau in artificial blood serum. After the exfoliation of HBN, its surface was coated with polydopamine (PDA) in alkaline conditions. The Anti-T-Tau was immobilized on a hydrophilic nanocomposite surface using PDA's reactive catechol and quinone groups, eliminating the need for extra crosslinkers. The working electrode surface of the screen-printed carbon electrode (SPCE) was coated with HBN-PDA nanocomposite using the drop-casting method. The biofunctional surface was created by directly immobilizing Anti-T-Tau on the HBN-PDA nanocomposite-modified SPCE. The analytical performance of the HBN-PDA/Anti-T-Tau/T-Tau immunosensor in the presence of T-Tau isoforms was determined through electrochemical measurements. The linear detection range was 1–30 pg/mL with a detection limit of 0.42 pg/mL for T-Tau, which is suitable for detecting T-Tau in the blood serum.