Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic with over 5 million fatalities. Vaccines against this virus have been globally administered; however, SARS-CoV-2 variants with spike protein mutations are continuously identified with strong capability to escape vaccine-elicited protection. Due to the high mutation rate and transmission ability, the development of a broad-spectrum SARS-CoV-2 inhibitor is highly in demand. In this study, the effect of poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS) against SARS-CoV-2 was investigated. The treatment of pseudoviruses carrying the SARS-CoV-2 spike protein with PEDOT:PSS strongly blocked SARS-CoV-2 pseudovirus infection in human ACE2-expressing cells without causing cytotoxicity. Specifically, PEDOT:PSS showed great potential in both inactivating viruses and rendering antiviral activity to the treated cells. The effects of other PEDOT:PSS solutions with different chemical ratios and properties were also validated to find the high inhibition capacity against SARS-CoV-2 pseudovirus infection. The transcriptomic data reveal that PEDOT:PSS-treated cells were endowed with transcriptional alteration, and it could be reverted after the removal of PEDOT:PSS from the culture medium. Importantly, PEDOT:PSS also exhibited broad-spectrum inhibition effects on the pseudovirus carrying the spike protein isolated from different variants. In combination with the advantage of high biocompatibility, PEDOT:PSS could thus be considered a potential therapeutic and prophylactic material against SARS-CoV-2.