Mitochondrial content estimated by MTG was found slightly but significantly increased in both control and Rapamycin 53123-88-9 patients cells in GAL medium. AICAR induced a clear increase in mitochondrial content only in the patient’s cells. The alteration of TMRE stain relative to mitochondrial content in the presence of AICAR was minor and not significant, indicating that Dy was not substantially affected. While uncoupled oxygen consumption was increased in GAL medium, AICAR had no significant effect although the basal uncoupled OCR was somewhat, not significantly relative to basal OCR in the patient’s cells. To examine to the downstream effect of AICAR, we performed immunostaining with an antibody towards Thr172 phosphorylated AMPK while co-staining with MTR. While clearly present in control cells, pAMPK stain was very weak in NDUFS2 cells grown on GLU. AICAR supplementation caused a marked and significant increase of pAMPK in the patients cells. Mitotracker stain was increased in patient’s cells on GAL but approached control values in the presence of AICAR. The search for therapeutic agents for mitochondrial complex I deficiency and OXPHOS defects in general, is seriously hampered by the lack of a standardized model system for evaluating treatment. Many studies focused on small groups of patients simultaneously treated with several agents leading to difficulties in interpreting data and patient responses to therapy in vivo. Documented in vitro assays usually focus on a specific compound or a specific parameter using a relatively large sample size. We developed an accessible and relatively simple system in 96 well plates assessing a number of different parameters by the use of one instrument. This enabled us screen multiple compounds on a small amount of fibroblasts simultaneously while measuring a number of different parameters. The small sample size allowed the use of primary cells which is advantageous not only for practical reasons but also because immortalized cells frequently do not retain their original phenotype and thus respond differently than primary cells. The fibroblasts for this study were chosen to represent CI deficiency, the most common OXPHOS defect. The cells were derived from patients with different CI defects, in order to assess individual responses to different compounds. Indeed the responses differed in some instances between the patients. This is exemplified by bezafibrate which was beneficial for NDUFS2 ATP content but not for C20ORF7, emphasizing the need to evaluate compounds on an individual basis. Individual evaluation is especially important when attempting to treat disorders where the mitochondrial function is already a priori compromised. The necessity to measure different parameters was evident when observing the effect of the various polyphenolic cytochemicals included in the study. Generally they decreased ROS formation which is advantageous, but concomitantly decreased growth and ATP content. Although some of these results contradict studies reporting positive effects previously mentioned in the introduction, they are actually in accord with other studies reporting that some polyphenols can induce cancer cell death.