The COVID-19 pandemic has revealed vulnerabilities in healthcare systems worldwide, emphasizing the importance of healthcare worker safety through adequate utilization of personal protective equipment (PPE). This study aims to assess the impact of pre-pandemic PPE training on the practices and other associated factors among frontline healthcare workers during the COVID-19 pandemic in Pakistan and provide insights into the implications of such training programs for future initiatives. A cross-sectional study from May 9th to June 5th, 2020 was conducted among the frontline healthcare workers against COVID-19 in Pakistan, utilizing an online structured questionnaire shared via WhatsApp and Facebook by using purposive sampling. Statistical analyses, including chi-square tests for proportion and logistic regression for the association while multi-logistic regression for potential confounders, were performed using SPSS version 22. A total of 453 healthcare staff participated, with 68.9% (n = 312) reporting no prior PPE training and 31.1% (n = 141) having received training. Significant associations were found between prior training and healthcare group distribution (p = 0.006), with doctors exhibiting the highest proportion of training 82 (37.61%), followed by nurses 50 (27.32%) and paramedics 9 (17.31%). Those who didn't receive any prior training in PPEs showed a higher perceived professional risk of 216 (69.23%) compared to those who received prior PPE training 96 (30.77%, p-value 0.005). Similarly, a higher frequency 137 (63.72%) of Perceived Personal risk was observed in those who didn't receive training, labeled as "high risk" compared to those who were trained 78 (36.28%, P value 0.02). Multi-logistic regression analysis identified paramedics as 0.26 times less likely to have received prior PPE training (Adjusted OR 0.26, 95% CI 0.10-0.65, p = 0.01) compared to medical doctors. Healthcare workers in tertiary care hospitals were 0.46 times less likely to undergo PPE training (Adjusted OR 0.46, 95% CI 0.25-0.87,p = 0.01) compared to those working at COVID-19 facilities/hospitals/quarantine centers. Likewise, individuals who doffed disposable gowns [Adjusted OR 3.86, (95% CI, 1.23-12.08, p = 0.02] were 3.86 times more interested in getting prior training in PPE compared to those who don't have skills to wear them. Our findings highlight that healthcare levels, type of healthcare, and doffing skills are important predictors of whether healthcare workers have taken prior training in PPE. These findings imply developing effective training programs for healthcare workers to ensure safety while providing care during pandemics like COVID-19.