The emergence of SARS-CoV-2, the virus causing COVID-19, has resulted in a pandemic that has disrupted all sectors of society. Less than a year after the sequencing of the virus’s genome, emergency use authorization for the BNT162b2 vaccine was requested. The aim of this study was to evaluate the response to the BNT162b2-mRNA COVID-19 vaccine in frontline workers from two hospitals in Colombia. Nasopharyngeal swabs were collected for the molecular detection of SARS-CoV-2 using real-time PCR, and blood samples were taken to assess seroconversion using qualitative and quantitative IgA, IgG, and IgM test kits. The study was conducted at a high-complexity healthcare institution in Bucaramanga, Colombia. 245 people were included in the first round and 129 in the second. SARS-CoV-2 molecular tests were conducted by RT-qPCR, and peripheral blood samples were collected to measure IgG, IgM, and IgA. The main outcome was to establish natural infection and the antibody response induced by the vaccine. The entire population was tested at two fixed times with RT-PCR tests and antibody level measurements approximately 4 and 8 months after receiving the second vaccine dose. 62 (25.3%) and 35 (14.3%) participants had a history of positive PCR in the first and second rounds, respectively. All positive cases showed elevated levels of all immunoglobulins, especially IgG. The average concentrations of IgA, IgM, and IgG at 90 days were 1149.5 U/mL (95% CI 828.2-1470.9); 320.3 U/mL (95% CI 218.4-422.3); and 9277.3 U/mL (95% CI 8989.2-9565.3). Frontline healthcare workers showed an adequate response to the BNT162b2 mRNA vaccine.