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Objectives: Metabolic dysfunction‐associated steatotic liver disease (MASLD) is the most prevalent liver disorder in Western countries, with approximately 20%–30% of the MASLD patients progressing to severe stages. There is an urgent need for noninvasive, cost‐effective, widely accessible, and precise biomarkers to evaluate liver steatosis. This study aims to assess and compare the diagnostic performance of a novel reference frequency method‐based ultrasound attenuation coefficient estimation (ACE) in both fundamental (RFM‐ACE‐FI) and harmonic (RFM‐ACE‐HI) imaging for detecting and grading liver steatosis. Methods: An Institutional Review Board‐approved prospective study was carried out between December 2018 and October 2022. A total number of 130 subjects were enrolled in the study. The correlation between RFM‐ACE‐HI values and magnetic resonance imaging proton density fat fraction (MRI‐PDFF), as well as between RFM‐ACE‐FI values and MRI‐PDFF were calculated. The diagnostic performance of RFM‐ACE‐FI and RFM‐ACE‐HI was evaluated using receiver operating characteristic (ROC) curve analysis, as compared to MRI‐PDFF. The reproducibility of RFM‐ACE‐HI was assessed by interobserver agreement between two sonographers. Results: A strong correlation was observed between RFM‐ACE‐HI and MRI‐PDFF, with R = 0.88 (95% confidence interval [CI]: 0.83–0.92; P <.001), while the correlation between RFM‐ACE‐FI and MRI‐PDFF was R = 0.65 (95% CI: 0.50–0.76; P <.001). The area under the ROC (AUROC) curve for RFM‐ACE‐HI in staging liver steatosis grades of S ≥ 1 and S ≥ 2 was 0.97 (95% CI: 0.91–0.99; P <.001) and 0.98 (95% CI: 0.93–1.00; P <.001), respectively, and 0.76 (95% CI: 0.65–0.85) and 0.80 (95% CI: 0.70–0.88) for RFM‐ACE‐FI, respectively. Great reproducibility was achieved for RFM‐ACE‐HI, with an interobserver agreement of R = 0.97 (95% CI: 0.94–0.99; P <.001). Conclusions: The novel RFM‐ACE‐HI method offered high liver steatosis diagnostic accuracy and reproducibility, which has important clinical implications for early disease intervention and treatment evaluation. [ABSTRACT FROM AUTHOR]