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Abstract: Objective To investigate the application of cytopathic effect (EMA)-PCR to rapid identification of infectious and non-infectious adenovirus. Methods The different dilutions of virus were inoculated to well-grown monolayer cells and then calculated the concentration of virus through the cytopathic effect (CPE). The adenovirus and New Castle disease virus were prepared in a fixed concentration of 10⁶ PFU/mL then diluted in ten-time gradient and the DNA of virus were extracted and PCR were applied, then the target DNA fragments were observed through gel electrophoresis. The non-infectious adenovirus were treated with different concentrations of EMA (0 μg/mL, 70 μg/mL, 120 μg/mL and 150 μg/mL) and then detected the target DNA fragment in the same way. The different concentrations of infectious adenovirus (10⁷ PFU/mL, 10⁶ PFU/mL, 10⁵ PFU/mL, 10⁴ PFU/mL and 10³ PFU/mL) were treated with 120 μg/mL EMA and then detected target DNA fragment in the same way. Results The target DNA fragments were detected in adenovirus of 10⁴ PFU/mL and above. The target DNA fragments were detected in all 3 types of adenovirus while not detected in New Castle disease virus. The DNA amplification of non-infectious adenovirus were suppressed by EMA at 120 μg/mL and 150 μg/mL concentration, while the DNA amplification of infectious adenovirus in concentration of 10⁵ PFU/mL and above were not affected by 120 μg/mL EMA. Conclusion EMA-PCR was proved to be feasible to identify the infectious and non-infectious adenovirus, in which false positive results of PCR were effectively avoid at the same time.