The amplitude of low frequency fluctuation (ALFF) is usually computed from T2*-weighted functional MRI (fMRI) signals, which is affected by R2* fluctuation, echo time, spin density, and mean R2*. To make ALFF more comparable and reliable between researches, we proposed a new method of ALFF calculation based on R2* (ALFF-R2*) in this work and further implemented for multi-echo fMRI. Statistical results from eyes-open and eyes-closed fMRI datasets show that ALFF-R2* detected an overall smaller significant regions than traditional ALFF in the paired T maps. The dice similarity coefficient between T maps from ALFF-R2* and traditional ALFF was 0.37. ALFF-R2* is potential to provide more specific results than traditional ALFF.