Autoimmune rheumatoid arthritis (RA) is a systemic condition closely correlated with a variety of autoantibodies (Abs) that could be considered diagnostic and prognostic markers. The current research was designed to detect the diagnostic values for a number (n) of these auto-Abs in RA detection and to evaluate the accuracy of a combined diagnostic scheme. This prospective study was conducted between September 2021 and August 2022 and included 110 subjects with RA, 70 individuals with other autoimmune disorders as positive controls (PC), and 50 unrelated, apparently healthy individuals as healthy controls (HC). The eligibility criteria for all study groups were followed stringently. An enzyme-linked immunosorbent assay (ELISA) was employed to measure rheumatoid factors (RF), cyclic citrullinated peptide antibodies (CCP-Abs), mutated citrullinated vimentin antibodies (MCV-Abs), anti-perinuclear factor antibodies (APF-Abs), and anti-keratin antibodies (AKA). We calculated the specificity, sensitivity, and predictive values of all auto-Abs. Significantly higher levels of anti-CCP-Abs, anti-MCV-Abs, APF-Abs, and AKAs were reported in the RA patients compared to the HC and PC subjects. RF levels, however, were only statistically elevated when compared to the HC individuals. Anti-APF-Abs had a higher sensitivity rate (70.9%), and anti-CCP-Abs had a higher specificity rate (94.16%) compared to other auto-Abs, whereas the combined detection scheme revealed a higher sensitivity (81.81%) and excellent specificity (90.83%) compared to the two former auto-Abs. Anti-perinuclear factor-Ab was a highly sensitive test, and CCP-Ab was a surpassingly specific assay for identifying RA. Furthermore, the combined detection scheme is an essential serological approach for RA diagnosis and crucial in differentiating this disease from other autoimmune diseases, thus promoting early diagnosis and treatment.