Multi-Method LCMS Assay Multiplexing with Advanced Analytical Intelligence Capabilities to Increase Laboratory Throughput

Importance of the Topic

High-throughput analysis is critical in clinical, forensic and industrial laboratories. Multiplexed LC-MS/MS approaches reduce instrument downtime, streamline workflows and cut per-sample costs. By running multiple assays concurrently on a single platform, laboratories can accelerate data generation and improve resource utilization.

Objectives and Study Overview

This study demonstrates the performance of the Shimadzu Nexera QX multiplexing LCMS system coupled to an 8060NX triple quadrupole mass spectrometer. Two distinct assays—benzodiazepines and synthetic cathinone (“bath salts”)—were overlapped in a 4-stream configuration. Key goals included assessing quantitative accuracy, linearity, carryover and reproducibility, as well as measuring time savings achieved by running methods in parallel.

Methodology and Instrumentation

The system comprised:

• Nexera QX four-channel multiplexing LCMS with dedicated pump for stream flushing
• Shimadzu LCMS-8060NX triple quadrupole MS
• QX Solution software for scheduling, AI-driven rinse and error recovery

Samples (urine extracts) were prepared via solid-phase extraction. Two analytical methods were configured:

• Benzodiazepines: 5.0 min run, 0.1% formic acid aqueous and methanolic phases, Kinetex Biphenyl column
• Bath salts: 2.5 min run, formic acid buffer and methanol with ammonium formate, Kinetex C18 column

Active needle and stream rinsing with four solvent selections and ternary gradient flushing minimized carryover while methods overlapped across streams.

Key Results and Discussion

Calibration linearity was excellent (R² ≥ 0.993) across both assays. Calibrator and QC accuracy ranged from 89% to 119%. Carryover was effectively suppressed by customized rinse protocols. Reproducibility over 1 000 injections showed <5% RSD in retention times and <10% RSD in internal standard areas. Multiplexing saved ~30 min per two 96-well plates compared to sequential single-method runs.

Benefits and Practical Applications

• Reduced cycle times and increased sample throughput
• Lower solvent consumption and minimized manual interventions
• Enhanced method flexibility for mixed-volume assay panels
• Suitable for clinical toxicology, forensic screening and QA/QC labs

Future Trends and Potential Applications

Advances may include expanded number of concurrent streams, integration with real-time AI for dynamic scheduling, coupling with high-resolution MS and full automation of sample preparation. Such developments will further boost throughput, reduce costs and support complex multi-analyte workflows in regulated environments.

Conclusion

The Nexera QX multiplexing system paired with QX Solution software enables robust, high-throughput analysis of multiple assays on a single LC-MS/MS platform. Excellent linearity, accuracy and reproducibility were achieved alongside significant time savings, demonstrating its value for laboratories seeking to maximize instrument utilization and operational efficiency.