Needle electromyography (EMG) of the tongue is traditionally used as a key to the diagnosis of amyotrophic lateral sclerosis (ALS), although relaxation of the tongue is often difficult to achieve. Recently, frequent abnormalities in the EMGs of the sternocleidomastoid (SCM) and upper trapezius muscles in ALS have been reported. To elucidate the diagnostic utility of these muscles we performed a multicenter prospective study to examine EMGs of the tongue (genioglossus), SCM, and trapezius in 104 ALS or suspected ALS patients. We also examined EMGs of the SCM and trapezius in 32 cervical spondylosis (CS) patients. We mainly evaluated fibrillation potentials/positive sharp waves (Fib/PSWs) and fasciculation potentials. Complete relaxation was achieved in 85% of ALS patients in the trapezius, but in only 6% of patients in the tongue. Fib/PSWs were observed in 8%, 13%, and 45% of ALS patients in the tongue, SCM, and trapezius, respectively, whereas fasciculation potentials were observed in 1%, 7%, and 39%, respectively. Abnormal spontaneous activity of any type was found in 9%, 17%, and 63% of patients, respectively. The high frequency of abnormal spontaneous activity in the trapezius was similar among the different diagnostic categories, and even 72% of clinically suspected ALS (progressive muscular atrophy) patients showed them in their trapezius. We did not observe Fib/PSWs or fasciculation potentials in any of our CS patients, thus these findings have excellent specificity. Tongue EMG added little utility over the clinical sign of tongue atrophy. Abnormal spontaneous activity in the trapezius would be more useful for the early diagnosis of ALS.