In vivo pressure-volume loops (PVLs) are the gold standard measurement to assess ventricular function. We developed a pipeline to integrate hemodynamic measurements with real-time three-dimensional (3D) echocardiographic data to construct in vivo PVLs for 25 post-heart transplant patients. We then evaluated left ventricular diastolic function for these patients by calculating chamber stiffness from a cubic polynomial fit of the diastolic pressure-volume relationships (PVR). We examined the ability of a well-established mathematical (Klotz) model to predict the patient-specific diastolic PVRs. We found that beat-to-beat variation in hemodynamic measurement was typical for this group of patients, which resulted in mean ± standard deviation end-diastolic chamber stiffness estimates of 0.75 ± 0.40 mmHg/ml. The cubic polynomial fits of the individual diastolic PVRs resulted in much smaller errors (0.25 ± 0.01 mmHg) compared to those associated with the Klotz predicted diastolic PVRs (4.0 ± 0.27 mmHg), which provided a poor representation of the in vivo diastolic PVRs. The proposed framework enables the temporal alignment between hemodynamic and 3D imaging data to produce in vivo PVLs that can be used not only to quantify global ventricular function, but also to estimate mechanical properties of the myocardium.