Cross-sectional area (CSA) is a fundamental variable in characterizing muscle mechanical properties. Typically, the CSA of a single muscle fibre is assessed by measuring either one or two diameters, and assuming the cross-section is either circular or elliptical in shape. However, fibre cross-sections have irregular shapes. The accuracy and precision of CSAs determined using circular and elliptical shape assumptions are unclear for mammalian skinned muscle fibres. Second harmonic generation imaging of skinned rabbit soleus fibres revealed that the circular assumption overstated real CSA by 5.3±25.9% whereas the elliptical assumption overstated real CSA by 2.8±6.9%. A preferred rotational alignment can bias the circular assumption, as real CSA was overstated by 22.1±24.8% when using the larger fibre diameter and understated by 11.4±13% when using the smaller fibre diameter. With 73% lower variable error and reduced bias, the elliptical assumption is superior to the circular assumption when assessing the CSA of skinned mammalian fibres.
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