Bronchoalveolar lavage (BAL) and protected specimen brushing (PSB) are the most commonly used methods for diagnosing ventilator-associated (VA) pneumonia although they require bronchoscopy. Endotracheal aspiration (EA) is a simple and less costly technique than PSB or BAL. The purpose of our study was to investigate the diagnostic value of EA quantitative cultures and to compare the results obtained using EA with those obtained using PSB and BAL in mechanically ventilated patients with or without pneumonia. We prospectively studied 102 intubated patients divided into three diagnostic categories: Group I (definite pneumonia, n = 26), Group II (uncertain status, n = 48), and Group III (control group, n = 28). All patients received prior antibiotic treatment. EA, PSB, and BAL were obtained sequentially in all patients. When comparing Group I with Group III and using 105 cfu/ml as a threshold, we found that EA quantitative cultures represented a relatively sensitive (70%) and relatively specific (72%) method to diagnose VA pneumonia. The specificity of BAL and PSB (87% and 93%, respectively) was better than that of EA. The negative predictive value of EA cultures was higher (72%) when compared with that obtained using PSB (34%) (p < 0.05). EA quantitative cultures correlated with PSB and BAL quantitative cultures in patients with definite pneumonia. Although EA quantitative cultures are less specific than PSB and BAL for diagnosing VA pneumonia, our results suggest that the former approach may be used to treat these patients when bronchoscopic procedures are not available.