In conclusion, we have developed high-throughput methodologies for quantifying chloride/nitrate antiport activity using a lucigenin assay. Validation with six previously characterised anionophores demonstrated that the EC50 values obtained were consistent with literature reports. Notably, these methods significantly reduce the time required compared to traditional approaches, and the integration of the Opentrons robotic liquid handling system enhances efficiency while minimising user error, making our protocols accessible to researchers with varying levels of experience.