Amyloid fibril structures of peptides having the sequence of the #2041 region of β2-microglobulin (β2m20-41 andβ2m21-29) were investigated with FTIR microscope spectroscopy. pH dependence of the amide I band of the IR absorption spectra was analyzed. For β2m20-41, the β-sheet content at pH 6.0 was 55% (12.1 residues) and the β-sheet was located in the N21G29 and I35V37 regions, but at pH 2.5 it was 60% (13.1 residues) and located in the F22V27 and P32V37 regions. The two structures with different amounts of β-sheet were switched at pH 4.5, which was close to the pI of this sequence (ca. 4.3) rather than the pKa of the carboxyl groups (5.8 ± 0.3 and 3.3 ± 0.4), suggesting the importance of the total number of charges rather than protonation of specific residue(s). The spectral characteristics of the IR linear dichroism and 13C isotope labeling indicated the formation of parallel β-sheet structure in both regions, and the stacking direction was unaltered by pH change. Unexpectedly however, the IR spectra of the β 2m21-29 fibril demonstrated the simultaneous presence of parallel/anti-parallel β-sheets. The relevance of the pH dependence of the β2m20-41 fibril structure to that of β2m fibrils is discussed.