Background: Amyotrophic lateral sclerosis (ALS) is the most common adult onset motor neuron disease. Therapy for ALS is limited to two drugs with modest effects on slowing disease progression. The diverse clinical presentation in patients and molecular pathomechanisms in genetic models suggest that distinct pathways converge on motor neuron death in ALS. Therefore novel biomarkers that stratify ALS patients into subgroups would offer insight into disease mechanism, point to new drug targets and allow group-specific clinical trials. Objectives: Our hypothesis is that fibroblasts share molecular pathomechanisms with the affected cells in ALS. Our goal is to test patient fibroblasts as a source of biomarkers for stratification. To this end we have measured functional bioenergetic parameters in large sample sizes of ALS patients and controls. We have also measured metabolic markers by mass spectrometry. Furthermore, we have undertaken large scale morphometric analysis of patient lines stained by vital dyes and immunocytochemistry specific to disease relevant cellular organelles and proteins under baseline and stress conditions. Results: We found that ALS fibroblasts display increased bioenergetic fluxes while showing similar cellular ATP levels as the controls. We were able to distinguish patients with upper motor neuron symptoms (PLS) from controls with high specificity and sensitivity based on bioenergetics. We have found that a subgroup of ~25% sporadic ALS patients displayed increased flux through the trans-sulfuration pathway. Conclusion: Subgroups of fibroblasts from ALS patients display metabolic alterations, such as hypermetabolism and hyper trans-sulfuration. These phenotypes may indicate an abnormal demand for energy production and anti-oxidant defense, respectively. We postulate that metabolism in itself is not sufficient to stratify all ALS patients into tight subgroups and evaluate imaging based biomarkers under stress conditions.