The Spike (K417T, E484K, N501Y) (SARS-CoV-2) Pseudotyped Lentiviruses were produced with SARS-CoV-2 Variant Spike (Genbank Accession #QHD43416.1 with mutations K417T, E484K, and N501Y) as the envelope glycoproteins instead of the commonly used VSV-G. These pseudovirions contain the firefly luciferase gene driven by a CMV promoter, therefore, the spike-mediated cell entry can be measured via luciferase activity. The Spike (SARS-CoV-2, K417T, E484K, N501Y) pseudotyped lentivirus can be used to measure the activity of neutralizing antibody against SARS-CoV-2 K417T, E484K, N501Y variant in intact cells using a Biosafety Level 2 facility.