In the last years, antibodies have emerged as a promising new class of therapeutics, due to their combination of high specificity with long serum half‐life and low risk of side‐effects. Diabodies are a popular novel antibody format, consisting of two F_v domains connected with short linkers. Like IgG antibodies, they simultaneously bind two target proteins. However, they offer altered properties, given their smaller size and higher rigidity. In this study, we conducted the—to our knowledge—first molecular dynamics (MD) simulations of diabodies and find a surprisingly high conformational flexibility in the relative orientation of the two F_v domains. We observe rigidifying effects through the introduction of disulfide bonds in the F_v–F_v interface and characterize the effect of different disulfide bond locations on the conformation. Additionally, we compare V_H–V_L orientations and paratope dynamics between diabodies and an antigen binding fragment (Fab) of the same sequence. We find mostly consistent structures and dynamics, indicating similar antigen binding properties. The most significant differences can be found within the CDR‐H2 loop dynamics. Of all CDR loops, the CDR‐H2 is located closest to the artificial F_v–F_v interface. All examined diabodies show similar V_H–V_L orientations, F_v–F_v packing and CDR loop conformations. However, the variant with a P14C‐K64C disulfide bond differs most from the Fab in our measures, including the CDR‐H3 loop conformational ensemble. This suggests altered antigen binding properties and underlines the need for careful validation of the disulfide bond locations in diabodies.