Background: The education and training of healthcare workers (HCWs) about pandemic response plans, infection control protocols (ICP), and precautions are critical for breaking the virus transmission chain. This study aimed to assess the knowledge of HCWs regarding the COVID-19 pandemic response plan and the ICP. Concomitantly, evaluating the effectiveness and impact of institutional measures, policies, and training programs. Methods: A cross-sectional study using an electronic questionnaire was used to collect data from 267 HCWs at a University-affiliated-hospital during April 2020. Results: In this study, 90.7% of participants were nurses and 66% were females. 60.3% of participants received specific training about caring for COVID-19 patients while at least 94.3% reviewed the pandemic response plan and ICP for COVID-19. 47.5% and 29.2% of participants provided direct care for suspected or confirmed COVID-19 patients, respectively. Between 85.7% and 100% of the participants answered all items correctly regarding the COVID-19 pandemic response plan, measures, and precautions. Furthermore, the correct answers were above 95% for each item. Conclusion: HCWs have a high level of knowledge about the COVID-19 pandemic response plan and ICP. Training and disseminating updated plans and protocols in different methods such as an intranet and onsite training in addition to regular methods were very effective.