Background: The purpose of this study was to compare different invasive methods for Helicobacter pylori ( H. pylori) detection, namely PCR for H. pylori specific ureC gene, Rapid urease test (RUT), and histopathological examination by modified Giemsa staining. Methodology: Endoscopic gastroduodenal biopsy materials were collected from dyspeptic patients who underwent endoscopic examination upon fulfilling the inclusion criteria. Three to four samples were collected from each patient after taking informed consent and proper clinical history. A rapid urease test (RUT) was done on spot with in-house RUT media from 1 specimen. One to two specimens were preserved in 10% formaldehyde for histopathology and PCR for ureC gene was done from 1 specimen. Collected biopsy specimens from gastric and duodenal mucosa of 142 patients were categorized as H. pylori-positive cases and H. pylori-negative cases based on the case definition used in the study upon positivity of 3 diagnostic tests. Results: Among 142 biopsy specimens, 34.5% were categorized as H. pylori-positive cases, 35.2% as H. pylori-negative cases, and finally 30.2% as doubtful or indeterminate cases. Rapid urease test was the most sensitive method, closely followed by ureC gene PCR and histopathology, with a sensitivity of 94.2%, 83.0%, and 76.5%, respectively. Whereas histology was the most specific, having 98.0% specificity followed by 83.0% in PCR. RUT was the least specific, with 55.5% specificity. Conclusion: While histopathology could detect H. pylori infection with the highest specificity, for definitive diagnosis combination of any 2 methods should be used, if available.