Amyotrophic lateral sclerosis (ALS) is a progressive disease of both upper motor neurons (UMNs) and lower motor neurons (LMNs) leading invariably to a decline in motor function. The clinical exam is foundational to the diagnosis of the disease and ordinal severity scales are used to track its progression. However, the lack of objective biomarkers of disease classification and progression delay clinical trial enrollment and muddle inclusion criteria. Ultimately, biomarker evidence of therapeutic target engagement will support and perhaps supplant more traditional clinical trial outcome measures. Electrophysiology tools of nerve conduction study and electromyography have already been established as diagnostic biomarkers of LMN degeneration in ALS, with the more recent introduction of motor unit number estimation (including MScanFit), motor unit number index, and electrical impedance myography. Dysfunction of UMN and non-motor brain areas is being increasingly assessed with transcranial magnetic stimulation, high-density electroencephalography, and magnetoencephalography. Although most of these techniques are used to explore the underlying disease mechanisms of ALS in research settings primarily, they have the potential on a broader scale to noninvasively identify disease subtypes, predict progression rates, and assess physiologic engagement of experimental therapies.