The invention relates to a an isolation debugging system of a transformer substation monitoring system, which comprises an in-operation monitoring subsystem and an isolation debugging subsystem, wherein the isolation debugging subsystem comprises a debugging server and a data transmission medium; and data interaction is carried out between the isolation debugging subsystem and the in-operation monitoring subsystem through the data transmission medium. The enlargement process of a transformer substation does not influence the normal operation of the original monitoring system according to the technical scheme of the invention, and a reformed and used server network and a network of the original monitoring system are relatively dependent and do not influence each other. The normal monitoring operation of the monitoring system to original operating equipment can not be influenced, and mistake operations to the original operating equipment through the monitoring system due to certain human factors can even be avoided.