<p>Methods based on photo-heat conversion materials have been developed for the immunoassays, but most involve low sensitivity or complex procedures and are unsuitable for routine use. Herein, a portable photothermal immunoassay was designed for point-of-care (POC) testing of disease-related biomarker (carcinoembryonic antigen (CEA)) in biological fluids on a digital near-infrared (NIR) thermometer with smartphone readout. Initially, the photothermal materials with manganese (Mn) and iron (Fe) bimetallic oxide (MnFe₂O₄) were synthesized via a typical hydrothermal method, and then functionalized with polyclonal anti-human CEA detection antibody (pAb₂). In the presence of target CEA, a sandwich-type immunoreaction was implemented on monoclonal rabbit anti-human CEA capture antibody-coated microplate with MnFe₂O₄-pAb₂. Under an 808-nm laser irradiation, the labeled MnFe₂O₄ on the detection antibody converted the light into heat, thereby causing the increasing temperature of the detection solution in the microplate. The variation in the temperature was collected on a portable NIR thermometer with smartphone readout. Under optimal conditions, the&#xa0;MnFe₂O₄-based immunoassay exhibited good photothermal responses for the detection of target CEA with a dynamic linear range of 0.01–150&#xa0;ng&#xa0;mL⁻¹ at a low detection limit of 0.56&#xa0;pg&#xa0;mL⁻¹. Good reproducibility, intermediate precision, and high specificity were acquired. No significant differences at the 0.05 significance level were encountered for the analysis of human serum samples between the developed photothermal immunoassay and the commercially available enzyme-linked immunosorbent assay (ELISA) for the determination of CEA. Importantly, this immunoassay offers promise for simple and cost-effective analysis of biological samples in diagnostics.</p> Graphical Abstract <p></p>