Ventilator associated pneumonia (VAP) is one of the most common healthcare associated infections in critical care settings. VAP is most commonly caused by biofilm producing organisms. Objectives: This study aimed to investigate biofilm formation on the luminal and surface of ETTs of children in ICU on mechanical ventilation and study its relation to VAP. The presence of biofilm on the surface of ETTs and biofilm stage were evaluated by Scanning Electron Microscope (SEM), meanwhile, bacteria harvested from the surface of ETTs and the secretions of lower respiratory tract were isolated, identified and assessed for antimicrobial susceptibility. Methods: Twenty ETTs from 20 child patients who were intubated and mechanically ventilated in the ICU of pediatrics hospital Cairo university during the period from November 2012 to May 2013 Results: The results revealed that Bacterial colonization and biofilm formation was observed on 85% of inner surface of collected ETTs using SEM. Staging of biofilm revealed that 64.7% of the biofilms were grade III, 23.5% were grade II, and 11.7% were grade I. Bacterial colonization and biofilm grading was shown to be time dependent (p=0.0005).Inner endotracheal tube surface yielded 70% positive aerobic cultures. The most prevalent isolates were Klebsiella pneumoniae followed by coagulase negative Staph (CoNS), Acinetobacter, pseudomonas aeruginosa and proteus mirabilis . Nine samples had the same pathogen both on the surface of ETTs and in the secretions of lower respiratory tract which accounted for 52% of the positive cultures from ETTs. All Gram negative isolates were resistant to, cefepime, cefotaxim, cefotriaxon, ampicillin-sulbactamIn conclusion : We can conclude that endotracheal tube colonization and biofilm formation are frequently observed in children undergoing mechanical ventilation, increases with the duration of intubation and is correlated with occurrence of VAP. Bacteria implicated in VAP showed multi-resistance towards most antibiotics used in the study.