Campylobacteriosis incidence and proportion of Campylobacter strains resistant to antibiotic has been increasing worldwide in the last decades. Campylobacteriosis is considered as one of the major important zoonotic gastrointestinal diseases around the world. The aim of this study is isolation and biochemical characterization of Campylobacter from poultry. Campylobaceter isolation have been done on MCCDA media, motility have been detected by phase contrast microscope , morphology have been detected by Gram stain, and confirmed by MALDI-TOF MS and molecular confirmation by using cPCR by amplification of 16SrRNA gene. A total of 102 isolate of Campylobacter were isolated from (farm and small backyards located in different sources in El-Kalyobia , El-Monofia and El-sharkia Governorate) by conventional bacteriological methods from which 40 were C.jejuni and 62 were C.coli. All isolats have been confirmed as Campylobacter by MALDI-TOF MS and detection of 16SrRNA were detected by cPCR in all isolates. PCR is a useful molecular tool for identification and confirmation of Campylobacter, its Rapid, sensitive and specific than the culture methods the only disadvantage is that is expensive.