We assessed humoral and cytokine responses in monthly plasma samples from ALS patients who received glatiramer acetate (GA) immunization every day or every other week, or remained untreated (control) from a six-month phase II trial. Samples were evaluated by GA-specific ELISA assays for detection of combined immunoglobulin (Ig) classes (IgM,A,G), IgG alone, and IgG subclasses (IgG1, IgG2, IgG3, and IgG4). T-helper (Th) type 1 and 2 (Th1 and Th2) cytokine levels were determined by flow cytometric cytokine bead arrays. Fourteen of 21 GA-immunized patients produced anti-GA Ig responses. Those treated every day produced anti-GA responses within one month, while those treated every other week exhibited responses by month two. All anti-GA IgG subclass concentrations were increased in excess of 4.2-fold in plasma from treated patients, and anti-GA IgG1 comprised the majority of the humoral response. Mean plasma cytokine levels were statistically indistinguishable between treatment regimens; however, stratification by patient and time on study showed more prevalent trends in changes of Th1 or Th2 cytokine levels following GA treatment every other week or every day, respectively. These data show significant humoral responses and cytokine trends following GA immunization in ALS patients.