Sheeppox virus (SPPV) is one of the listed and notifiable disease affect sheep production with major effect on the trade of new breed of sheep. This study was conducted to identify sheep pox by using cell culture, electron microscope (EM) and open reading frame (ORF) 103 gene during an outbreak of local breed of sheep occurred in Sharkia Governorate, Egypt in April 2017. Affected adult sheep showed typical skin pox lesion on face, the inner side of lips, inner aspect of the thigh and under the tail. The incidence rate of infection was 23.5% and the mortality rate in young lambs aged 3 to 6 months old was 8.2%. Forty-three scabs and tissue samples from clinically diseased adult sheep and dead lambs were collected and subjected to culture on Vero cell. The cytopathic effect (CPE) was observed within 3 to 7 days in 40 samples. A typical Poxvirus was a brick-like shape with round ends by negative staining of EM and ovoid like structure with dumb-bell shaped DNA core with concave bodies sides by positive staining of EM. By conventional PCR utilizing ORF103 gene and obtained bands of about 570 bp referred to SPPV. The sequence amplicon was analyzed by NCBI-BLAST and register in GeneBank under accession N. MG873537 and phylogentic tree was designed which revealed that the isolated strain of SPPV was resembled with other strains of SPPV isolated in Egypt, India, China, and USA. Finally, both EM and PCR are considered as sensitives, rapids, and powerful methods to identify SPPV from tissue and scab’s samples without the need of further culture in addition to the useful and easily use of ORF103 gene to differentiate SPPV from other Capripoxvirus.