The E/e' ratio has an established role in the assessment of left ventricular filling pressure (LVFP) in stable patients, but its accuracy in acute myocardial ischemia is less well established. The aim of this study was to validate the relation between the E/e' ratio and invasively measured LVFP in patients with non-ST elevation myocardial infarction (NSTEMI). A total of 120 unselected patients with NSTEMI underwent cardiac catheterization with measurement of left ventricular end-diastolic pressure (LVEDP; elevated ≥15 mm Hg) and Doppler echocardiography with either simultaneous (n = 30) or same-day (n = 90) measurement of E/e'. Patients were aged 64.1 ± 11.8 years, 72% were male and mean left ventricular ejection fraction was 48.0 ± 20.9%. Septal, lateral, and average E/e' ratios all showed a significant correlation with LVEDP (Pearson's r: 0.42, 0.43, 0.48, respectively [all p <0.001]). Receiver operating characteristics curves showed an area under the curve of 0.72, 0.72, and 0.75 (all p <0.001) for septal, lateral, and average E/e', respectively. The sensitivity, specificity, positive (PPV), and negative (NPV) predictive values for the guideline-recommended threshold of average E/e' >14 for elevated LVEDP was 27%, 93%, 79%, and 44%, respectively. Utilizing lower E/e' boundaries of 6, 7, and 8 for lateral, average, and medial E/e', respectively, improved the NPV to ≥80% for each parameter. In conclusion, the E/e' ratio is a robust measure of LVFP during acute NSTEMI using upper and lower thresholds to achieve a high PPV and NPV, respectively, with the use of adjunctive guideline-recommend measures required in patients with nonconclusive E/e'.