The M1 matrix protein of influenza A virus, which plays multiple roles in virion assembly and infection, underlies the viral envelope. However, previous studies have given differing accounts of the number of layers in the M1/envelope complex and their thicknesses and compositions. To resolve this issue, we performed cryo-electron microscopy and cryo-electron tomography on the self-same specimens. At neutral pH, there are two kinds of complexes, corresponding to a lipid bilayer with embedded glycoproteins, with and without a closely associated, 4 nm-thick, sheet of M1 protein. The reported discrepancies arose from differences in imaging conditions; i.e. in defocus and in whole-particle projections vs. thin tomographic slices. Exposure of virions to low pH (as in the endosome) promotes membrane fusion and previous work has shown that the M2 ion channel causes the virion interior to acidify also. We found that after 5 min at pH 4.9, the proportion of virions lacking an M1 layer increases from 10% to 50%. In pH 4.9/5 min virions retaining an M1 layer, the M1/envelope complex exhibits two states: viz., the original, neutral pH state and one in which the M1 layer appears thinner and/or closer to the membrane. These observations extend previous indications that acidic pH causes the M1 layer to dissociate, leaving the envelope more pliable and consequently fusion-compatible, and show that dissociation is preceded by a conformational change in M1.