Simple SummaryA microfluidic biosensor based on a thin-core microfiber coated with molybdenum disulfide for immune detection ofToxoplasma gondiiwas illustrated. The detection range, limit of detection, detection sensitivity, specificity, and clinical properties of the proposed biosensor were explored.AbstractToxoplasma gondii(T. gondii) is a zoonotic parasite that is widely distributed and seriously endangers public health and human health. Therefore, accurate and effective detection ofT. gondiiis crucial. This study proposes a microfluidic biosensor using a thin-core microfiber (TCMF) coated with molybdenum disulfide (MoS2) for immune detection ofT. gondii. The single-mode fiber was fused with the thin-core fiber, and the TCMF was obtained by arc discharging and flame heating. In order to avoid interference and protect the sensing structure, the TCMF was encapsulated in the microfluidic chip. MoS2andT. gondiiantigen were modified on the surface of TCMF for the immune detection ofT. gondii. Experimental results showed that the detection range of the proposed biosensor forT. gondiimonoclonal antibody solutions was 1 pg/mL to 10 ng/mL with sensitivity of 3.358 nm/log(mg/mL); the detection of limit was calculated to be 87 fg/mL through the Langmuir model; the dissociation constant and the affinity constant were calculated to be about 5.79 × 10−13M and 1.727 × 1014M−1, respectively. The specificity and clinical characteristics of the biosensor was explored. The rabies virus, pseudorabies virus, andT. gondiiserum were used to confirm the excellent specificity and clinical characteristics of the biosensor, indicating that the proposed biosensor has great application potential in the biomedical field.