The invention discloses a method for detecting a neutralizing antibody in coxsackievirus A6 (CV-A6), and a recombinant virus applied in the method. According to the method, the neutralizing antibody is detected by using a pseudovirus system packaged by two types of recombinant plasmids; by adopting a single-cycle infected pseudovirus, the safety problem caused by use of live viruses is avoided. The results of a plurality of tests prove that the pseudovirus detection system is a CV-A6 neutralizing antibody detection method which is safe, sensitive, rapid, specific, simple and convenient, and low in cost. Based on the characteristics, the pseudovirus detection system is very suitable for the tests for rapid and large-scale detection of the neutralizing antibody, and has an import applicationvalue for development of a CV-A6 viral vaccine and detection of CV-A6 specific neutralizing antibody level of patients and populations.