Nonsense-mediated mRNA decay (NMD) is a eukaryotic quality control mechanism for the dynamic regulation of gene expression. NMD degrades transcripts containing a premature termination codon (PTC) more than 50-55 nucleotides upstream of the final exon-exon junction. Although NMD is a ubiquitous mechanism for degrading RNA transcripts in all eukaryotes, there is great variety in the efficiency and specificity of the degradation mechanism. While most transcripts containing a PTC are degraded via NMD, transcripts containing a PTC can evade NMD and produce truncated or full-length proteins1. NMD efficiency may also vary based on gene sequence, intracellular location, tissue, or on an individual level. This study aimed to aid the understanding of NMD as an endogenous control for gene expression by evaluating NMD efficiency in homogenous cell cultures. We evaluated NMD efficiencies in human embryonic kidney cells by transfecting cell cultures with dual-fluorescing reporters for NMD. We measured fluorescent levels through flow cytometry, and surprisingly detected varying NMD efficiencies among cells of the same culture. To investigate the possible causes of the range in NMD efficiency, we sorted cell cultures based on NMD efficiency levels and evaluated cell populations for their concentrations of NMD factors through immunoblotting and RT-qPCR. Results revealed that NMD factor expression levels did not correlate with NMD efficiency, which proposes new questions for the role of NMD factors in NMD and other possible intracellular mechanisms affecting NMD efficiency. We hypothesized that cell cycle may be affecting NMD. To study the possible relationship, groups of cells with varying NMD efficiencies were evaluated through immunoblotting for cell cycle stage. Preliminary results did not indicate a relationship; however, the association must be further evaluated. Conclusively, we determined a range in NMD efficiency among individual cells in homogenous human embryonic kidney cell cultures. We aim to progress this research by determining key factors and mechanisms that may influence NMD efficiency. Implications for understanding the specificities of NMD activity are far-reaching in the medical field, as several severe human diseases, such as facioscapulohumeral muscular dystrophy, are strongly tied to NMD inhibition.