The rapid and sensitive detecting technique for subgroup J of avian leukosis viruses (ALVs-J) needs to be developed as soon as possible in order to not only reduce economic losses but also ensure the food safety. In this paper, a novel electrochemical immunosensor was built based on mesoporous graphitic carbon nitride (mpg-C3N4). Mpg-C3N4 was used as the sensor platform to bond with the primary antibodies (Ab(1)). The compound of thionine and mpg-C3N4 (Th-mpg-C3N4) was synthesized for the first time to serve as the electroactive probe as well as the carrier of secondary antibodies (Ab(2)). Compared to bulk g-C3N4, mpg-C3N4 possesses larger specific surface area, smaller electrochemical resistance and abundant active sites. So the proposed electrochemical immunosensor exhibited amplified detective signals which realized the sensitive detection of ALVs-J. Under the opitimized conditions, the immunosensor exhibited outstanding analytical performance for the detection of ALVs-J whose titer ranged from 10(2.08) to 10(4.0) TCID50/mL (TCID50: 50% tissue culture infective dose) with a low detection limit of 120 TCID50/mL (S/N = 3). The sensitivity of the immnosensor was 6.15 mA/(TCID50/mL). This high sensitive immunosensor also displayed good selectivity, reproducibility and stability. Last but not least, this new strategy may be of great promise for clinic application in the future. (C) 2016 Elsevier Ltd. All rights reserved.