The RT-PCR assay for the detection of all human enteroviruses was used to analyze fecal samples, cerebrospinal fluid (CSF), respiratory tract swabs, and sera from 204 patients with symptoms of viral infection. Enteroviruses were detected in 31.37% of cases. The proposed method analyzes every type of clinical samples. The relative detection rate of enteroviruses in different samples was 70.64% in feces, 10.4% in SCF, 17.24% in swabs, and 21.14% in the sera. The use of serum in addition to other testing samples increased the rate of enterovirus detection at 18.25%. Developing techniques may be used for the diagnosis of acute enterovirus infections with different clinical manifestations.