Two doses of the BNT162b2 mRNA vaccine are highly effective against SARS-CoV-2. Here, we tested the antibody neutralization against Omicron SARS-CoV-2 after 2 and 3 doses of BNT162b2. Serum from vaccinated individuals was serially tested for its ability to neutralize wild-type SARS-CoV-2 (USA-WA1/2020) and an engineered USA-WA1/2020 bearing the Omicron spike glycoprotein. At 2 or 4 weeks post dose 2, the neutralization geometric mean titers (GMTs) against the wild-type and Omicron-spike viruses were 511 and 20, respectively; at 1 month post dose 3, the neutralization GMTs increased to 1,342 and 336; and at 4 months post dose 3, the neutralization GMTs decreased to 820 and 171. The data support a 3-dose vaccination strategy and provide a glimpse into the durability of the neutralization response against Omicron. Display omitted •Two doses of BNT162b2 are not sufficient to elicit robust neutralization against Omicron•Three doses of BNT162b2 confer substantial neutralization against Omicron•Neutralization against Omicron remains robust at 4 months after dose 3 of BNT162b2 Through serially testing BNT162b2-vaccinated human sera against SARS-CoV-2 and its Omicron variant, Xia et al. found that 2 doses of BNT162b2 are insufficient to elicit robust neutralization against Omicron. Three doses increase the magnitude and breadth of neutralization against Omicron and remains robust for up to 4 months post dose 3.