Throughput of the method
The LIFT method allows for a much greater measuring distance between the sensor and the target leaves than previous gas exchange and PAM-type ChlF measurement devices18,19, which in turn enables automated high-throughput measurements being conducted both in the field and in indoor environments12,32,37. This eliminates the need for physical interaction with the canopy. The throughput of the method increases with robot driving speed. To ensure accurate measurement of ChlF yield over time, the distance travelled during one Fq'/Fm' measurement must remain negligible compared to the illuminated area diameter (20 mm for the LIFT sensor). When the robot moves during excitation, the illuminated spot also shifts, so that ChlF yield per flashlet decreases, ultimately causing erroneous estimates of Fm' and Fq'/Fm' at excessive speeds. At a robot speed of 0.5 m s-1, only about 2.4% of the area illuminated by the last excitation flashlet was not already illuminated by the first excitation flashlet, meaning that the systematic measurement error on Fm' due to the motion of the robot will be numerically negligible (full calculation on GitHub, https://github.com/beat2keller/lift_data_processing). At a velocity of 0.5 m s-1, we measured 300 plots (1.5 × 2 m) per hour on a 40 × 36 m field, corresponding to several thousand plots per day. Moreover, the relatively low total weight (200 kg) and compact dimensions of our measuring setup allow transport by light vehicles, facilitating multi-site trials to assess photosynthetic efficiency across environments38.
Leaf versus canopy photosynthesis
The LIFT measurements in this method are spatially restricted to the uppermost, sunlit layer of the canopy, which is responsible for about 50% to 70% of total photosynthesis39. Furthermore, the amount of light each leaf can absorb critically depends on its angle relative to the sun, with more vertical leaves generally increasing leaf area index, enhancing light penetration and canopy photosynthesis40,41. With the LIFT sensor assembly used in this method, effects of 3D canopy architecture on single-leaf or whole-canopy photosynthesis can be modelled using the reflectance measurements from the spectrometer28 and/or an explicit reconstruction of 3D canopy geometry42 based on the images of the stereo RGB camera system. New algorithms, such as MASt3R36, learn dense, geometry-aware feature correspondences using deep transformers, enabling more robust and accurate 3D reconstruction across wide or repetitive regions. However, even without such additional corrections, the usefulness of automated, high-throughput methods, such as the LIFT, to identify more productive and resilient crop varieties has been demonstrated11,43.
Sink limitation
Sink limitation, i.e., active downregulation of photosynthesis due to an inability of the plant to use the amount of photosynthates it could produce44,45, can mask differences in εc among genotypes. Sink-limited downregulation of photosynthesis likely accentuates with increasing CO2 concentration in the atmosphere45. Our method can detect sink limitation only when it leads to a decrease also in Fq'/Fm'; there are indications that this can in fact occur46. In any case, the relevance of sink limitation appears to strongly depend on the crop and developmental stage, with wheat during grain filling being much more affected than grain legumes45 because the symbiotic N fixation of the latter constitutes a strong additional sink1,44,46. If sink limitation is expected to be of high relevance in a specific crop during the time period in which LIFT measurements are performed, further studies are needed to compare the resulting Fq'/Fm' values to simultaneously collected gas exchange data and check the validity of the assumption that Fq'/Fm' constitutes a good proxy for εc.
Measurement distance and variability in canopy height
The LIFT sensor assembly used in this method is designed to conduct measurements from a distance of approximately 60 cm to the crop canopy32,47. The LIFT sensor used in the method is fairly robust against minor differences in measuring distance, but a larger measuring distance generally appears to result in somewhat smaller values of Fq'/Fm'47. To prevent such bias, we recommend using the response of Fq'/Fm' to PPFR (which is more robust to changes in measurement distance) instead of the absolute values of Fq'/Fm' as in12 and / or to explicitly account for differences in canopy height during data analysis.
Unintended, artificial shading of leaves
Increased light intensity generally decreases Fq'/Fm'3,8,12. Thus, unnecessary shading by operators or the measuring setup should be avoided3,8,12. To minimize shading from the robot, the driving direction can either be set to prevent it entirely32 (at the cost of reduced throughput due to unproductive return trips), or measurements can be taken in alternating directions and then statistically corrected for shading effects. In this study, the term Heading × Hour within the statistical model of Fq'/Fm' (see step 4.3.10) ensures that shading of the target leaves by the robot is accounted for.
Conclusion
In summary, a LIFT sensor mounted on an autonomous robot enables fast and automated measurements of photosynthetic efficiency under field conditions, overcoming throughput limitations of previous approaches. Whereas factors such as canopy structure and sink limitation require careful consideration, our results demonstrate that reliable and scalable photosynthesis screenings in agronomic field trials and plant breeding nurseries are feasible. The ensemble of a high-resolution stereo RGB camera system and the LIFT as a point sensor will allow for a further increase in precision: AI-driven 3D localization of leaves will enable sampling leaves with similar orientation towards the sun and taking genotype-specific leaf inclination into account. From a research perspective, it may be particularly interesting to compare the photosynthesis of genotypes with varying canopy architecture and conduct high-throughput LIFT measurements under free-air CO2 enrichment (FACE) to better understand how Fq'/Fm' is affected by processes downstream of PS II under field conditions and potential sink limitations. In practical application, screening for photosynthetic performance and stress tolerance in breeding nurseries up to the farmer's field for precision farming applications has great potential.