When dealing with ill-posed problems such as fluorescence diffuse optical tomography (fDOT) the choice of the regularization parameter is extremely important for computing a reliable reconstruction. Several automatic methods for the selection of the regularization parameter have been introduced over the years and their performance depends on the particular inverse problem. Herein a U-curve-based algorithm for the selection of regularization parameter has been applied for the first time to fDOT. To increase the computational efficiency for large systems an interval of the regularization parameter is desirable. The U-curve provided a suitable selection of the regularization parameter in terms of Picard's condition, image resolution and image noise. Results are shown both on phantom and mouse data.