Here, we present an optimized enzyme-linked immunosorbent assay (ELISA) protocol for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike-specific immunoglobulin G (IgG) in plasma or serum samples. The protocol offers a rapid and reliable method, delivering results in less than 6 hours. On day 1, 96-well plates are coated with a concentration of 5 µg/mL of SARS-CoV-2 spike protein in 0.1 M sodium hydrogen carbonate and left to incubate overnight at 4 °C. Subsequently, the plates undergo thorough washing and are blocked with a buffer containing bovine serum albumin for one hour at room temperature. Diluted samples are added to the wells, followed by a two-hour incubation period. After another round of washing, a horseradish peroxidase-labeled antibody directed against the crystallizable fragment region of IgG is added, and the plates are incubated for one hour. The plates are then washed and developed using 3,3′,5,5′-tetramethylbenzidine-substrate for 30 minutes. The reaction is stopped by the addition of 1 N sulfuric acid. The optimized protocol demonstrated excellent performance, exhibiting a specificity of 99.5% and a sensitivity of 98.1% for the detection of SARS-CoV-2 spike-specific IgG in human plasma or serum samples.