Fig. 2. Categorical and quantitative comparisons.

a, Conventional objectives are constrained to a mechanical envelope that limits the product of FOV and WD. The general limits imposed by this mechanical envelope are sketched with dotted lines⁴³. The Cousa objective is distinctive in that it combines an ultra-long WD of 20 mm with a large FOV and an NA of 0.5. b, Conventional objectives that can offer a FOV >2 mm² are typically constrained to shorter WD. The Cousa has a long WD, and provides more than 4 mm² FOV. c, Overall throughput in both near IR (910 nm) and visible (532 nm) wavelengths is higher with the Cousa than conventional objectives. This is potentially due to a lower number of lenses in the Cousa. d, The FOV of the Cousa dwarfs both a commonly used short WD low magnification multiphoton objective (Nikon ×16/0.8 NA) and a conventional long WD objective (Mitutoyo ×20/0.4 NA). e, In vivo calcium dynamics (GCaMP6s) are measured with the Cousa objective and a popular water immersion objective (Nikon ×16/0.8 NA) and compared. The exact same imaging parameters are used to image the same field of neurons in the same mouse (awake, spontaneous activity). Averaged images for both objectives are shown on the left. Calcium traces from the identical ROI are shown on the right (raw data at 45 frames per s; no filtering). The data quality of the Cousa objective is similar to that of this commonly used short WD objective (Nikon ×16/0.8 NA). f, The signal-to-noise ratio and maximum ΔF/F of the calcium dynamic for each ROI in e are calculated and plotted as histograms for both objectives. The signal-to-noise ratio for each ROI is the ratio of the maximal magnitude of the calcium trace to the standard deviation of the fluctuating signal around the baseline. The maximum ΔF/F for each ROI is the maximal value of ΔF/F throughout the trace. Thus, the Cousa provides an ultra-long WD in air, a large FOV and raw data similar to those from conventional objectives.