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Purpose: To compare two automated semen quality analysis systems (LensHooke® X1 PRO [X1 PRO]) and IVOS CASA) for accuracy, precision and agreement with laboratory-based manual semen analysis (MSA). Materials and Materials and Methods:Methods: Semen samples (n=31) were obtained from normozoospermic healthy male volunteers and infertile men with a minimum abstinence period between 2–3 days. After complete liquefaction, 101 seminal aliquots were prepared and tested according to WHO 5th Edition (2010) guidelines. The results obtained by X1 PRO and IVOS CASA were com-pared with that of MSA. Additionally, 10 samples were used to evaluate the intra- and inter-rater agreement for X1 PRO and MSA. Results:Results: The semen parameters (sperm concentration, total, and progressive motility) showed strong correlation and agree-ment for both automated semen analyzers and MSA (Spearman’s rank correlation ≥0.92, p<0.0001). X1 PRO and IVOS CASA were able to differentiate samples with abnormal concentration with a positive predictive value (PPV) of 100%. Furthermore, the PPV for X1 PRO (86.5%) was higher than that for IVOS CASA (71.7%) in differentiating samples with abnormal motility. The X1 PRO device showed a high PPV (97.7%) in identifying normal sperm forms compared to MSA. Semen parameters evaluated showed a high inter-rater (kappa >0.91) and intra-rater (kappa >0.92) agreement for X1 PRO compared with MSA. Conclusions:Conclusions: Both automated semen analyzers demonstrated a high level of concordance and their performance was compa-rable with MSA analysis. Furthermore, high-levels of inter-and intra-rater reliability for semen analysis indicate that the new X1 PRO can be used in a clinical laboratory to offer accurate and quick test results.