Finding natural and/or synthetic ligands that activate orphan G protein-coupled receptors (oGPCRs) is a major focus in current drug discovery efforts. Transfluor is a cell-based GPCR screening platform that utilizes an arrestin-green fluorescent protein conjugate (arrestin-GFP) to detect ligand interactions with GPCRs. The assay is ideally suited for oGPCRs because binding of arrestin-GFP to activated receptors is independent of the interacting G protein. Before embarking on a high-throughput screen, it is important to know that the target oGPCR can actually bind arrestin-GFP. This information was thought to be inaccessible, however, as arrestin-GFP recruitment is an agonist-driven process. This chapter describes an assay that enables GPCRs to be validated in Transfluor in the absence of ligand. This assay, termed the ligand-independent translocation (LITe) assay, utilizes a modified G protein-coupled receptor kinase to bypass the requirement of ligand for initiating arrestin-GFP translocation. Using the LITe assay, one can determine if an oGPCR binds arrestin-GFP and if the response is quantifiable by high-content screening instruments. In addition, the assay expedites the development and identification of oGPCR stable cell lines with the best Transfluor properties. In this way, the assay provides criteria for selecting the best oGPCRs to move forward for a Transfluor screening campaign. Moreover, the assay can be used for quality control purposes during the orphan receptor screen itself by providing positive translocation responses for calculation of Z prime values. In summary, the LITe assay is a powerful new technology that enables a faster and more reliable path forward in the deorphanization of GPCRs with Transfluor.