Agilent ICP-MS Journal (May 2014 – Issue 57)

Význam tématu

The precise measurement of proteins in single cells and the rapid development of robust ICP-MS methods for trace-level elements are critical for advancing research in cell biology, disease mechanisms, food safety and environmental analysis. Techniques that combine high sensitivity, multiplexing capability and automated workflows address the growing demand for data quality and throughput in academic and industrial laboratories.

Objectives and Study Overview

Two complementary approaches are presented. The first describes a novel workflow for quantifying multiple proteins at the single-cell level by combining rare-earth-element immunolabeling, laser capture microdissection (LCM) and triple quadrupole ICP-MS (ICP-QQQ). The second demonstrates the Agilent MassHunter “Method Wizard” for the new 7900 ICP-MS, which automates method development and delivers accurate, routine analysis of certified reference materials (CRMs) for food matrices.

Methodology and Instrumentation

The single-cell protein assay uses antibodies tagged with 141Pr, 167Er, 171Yb and 103Rh to label myosin, IL-6, GAPDH and DNA. Individual muscle fibers are dissected by LCM, digested in nitric acid and analyzed by the Agilent 8800 ICP-QQQ under optimized MS/MS conditions. Carrier and dilution gases, RF power and sample introduction parameters are auto-tuned in MassHunter.

The 7900 ICP-MS study uses Agilent’s Method Wizard to guide users through automatic selection of plasma mode (Low Matrix, General Purpose or UHMI), cell gas (He, H2, high-energy He) and analyte isotopes based on a semi-quantitative scan of a “typical” CRM digest. Calibration and internal standard options are configured automatically to match sample composition. Three food CRMs (fish protein and two fish tissues) were digested by microwave and analyzed with UHMI and H2 cell gas options.

Key Results and Discussion

In single-cell samples, ICP-QQQ detected myosin (141Pr) and DNA (103Rh) with counts in the 1 000–3 000 range, and lower signals for IL-6 and GAPDH tags, yielding an average signal-to-noise ratio of 85. Fiber-to-fiber variability reflected biological heterogeneity. Control fibers without labels produced no detectable background.

For the food CRMs, concentrations of major (Na, K, Ca, Mg, P, S) and trace elements (V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Cd, Hg, Pb, U, Th) agreed with certified values within quoted uncertainties. The automated method required no user adjustments and delivered results with low bias and good precision.

Benefits and Practical Applications of the Method

• Single-cell multiplexing with ICP-QQQ enables highly sensitive, spatially resolved protein profiling in complex tissues.
• Automated method creation for the 7900 ICP-MS lowers the barrier to routine trace-element analysis and boosts laboratory productivity.
• Both workflows enhance data reliability, reduce development time and support QA/QC in life sciences, food safety, environmental and clinical testing.

Future Trends and Potential Applications

Emerging directions include integration of remote instrument monitoring via mobile apps, expansion of single-cell metal-tag assays to basic and translational research, and the extension of automated ICP-MS methods to environmental, pharmaceutical and geological matrices. Continuous improvements in cell-gas chemistry and high-matrix introduction will further increase method robustness.

Conclusion

The combination of rare-earth-element labeling, precise LCM sampling and ICP-QQQ offers a powerful platform for single-cell protein quantification. Meanwhile, automated method wizardry in the 7900 ICP-MS streamlines trace-element workflows, ensuring accurate results with minimal user intervention. Both approaches address key challenges in analytical chemistry and pave the way for broader adoption in diverse laboratories.

Used Instrumentation

• Agilent 8800 Triple Quadrupole ICP-MS with low-flow nebulizer and MS/MS mode.
• Leica LMD7000 laser capture microdissection microscope.
• Agilent 7900 ICP-MS with Ultra High Matrix Introduction and H2 cell gas.
• Agilent 1260 Capillary-LC and ASX-520 autosampler for sample introduction.
• Milestone ETHOS 1 microwave digestion system.

References

1. Wanagat J, Cao Z, Pathare P, Aiken JM (2001) Faseb J 15(2):322-332.
2. Agilent Application Note 5991-4257EN: High-salt matrix analysis by the Agilent 7900 ICP-MS with UHMI.
3. Agilent Application Note 5991-4556EN: Routine trace-metal determination in food CRMs using the 7900 ICP-MS Method Automation.