Background: Helicobacter pylori (H. pylori)colonizes the stomach and affect almost 50% of the world's population. Clarithromycin is considered a cornerstone for H. pylori treatment. Emergence of clarithromycin resistance (CLR-R) has played a major role in failure of H. pylori eradication both in adults and children.  Clarithromycin resistance is mostly due to mutations in 23S rRNA gene: A2142G, A2142C, and A2143G. The aim of the current study is to determine the prevalence of CLR-R among H. pylori infected children with prior clarithromycin treatment. Materials and Methods: Multiple endoscopic gastric biopsies were collected from 50 H. pylori infected children after cessation of clarithromycin-based treatment. Samples were subjected to histopathological examinations, rapid urease test (RUT) and simultaneous molecular detection of H. pylori infection as well as CLR-R by multiplex Real-Time polymerase chain reaction (PCR). Results: Histopathological examinations and RUT revealed H. pylori in 74% and 92% of samples respectively. Molecular detection of CLR-R showed that 62.5% positive H. pylori cases were not harboring any of the tested mutations, while 25% harbored 2143A-G single mutation. Double mutations (2142A-C and 2143A-G) were detected in only 4 cases. Statistical significant correlation existed between both RUT and PCR results as well as between histopathological findings and PCR test results. Conclusions: A combination of histopathogy, RUT and multiplex PCR procedures offers a real benefit in the simultaneous diagnosis of H. pylori infection along with clarithromycin resistance status. Other mechanisms of clarithromycin resistance need to be investigated to explain treatment failure in absence of the previously detected mutations.