Aflatoxins are poisonous secondary metabolites capable of causing toxicity in several food items and water. The routine monitoring of Aflatoxins has assumed great analytical significance. The electrochemical biosensors are attractive tools to screen the Aflatoxin-laden samples. The present study reports on a new type of electrochemical biosensor, based on an electrode prepared of multifunctional MOF (Metal-organic framework)/MXene composite material. Molybdenum-based 2-dimensional (2D) MOF and Ti3C2 MXene have been mixed to prepare the composite, which is then deposited over a screen-printed electrode (SPE). The anti-Aflatoxin B1 (AFB1) antibodies (Ab) are covalently complexed, and the prepared Ab-MOF/MXene sensor is investigated for its applicability to detect AFB1. Utilizing the electrochemical impedance spectroscopic (EIS) analysis, the Ab-MOF/MXene sensor has allowed the detection of AFB1 from 0.06 to 50 ng/mL with a very low limit of detection, i.e., 8 pg/mL. The sensor performs well with respect to all the major quality parameters, such as long-term stability, specificity in co-presence of interfering species, reproducibility, and repeatability. It is demonstrated successfully to detect AFB1 in the spiked samples of Pistachio.