Agilent ICP-MS Journal (May 2006 – Issue 27)

Significance of the topic

The emerging field of metallomics explores the distribution and role of metal ions in biological systems, a critical aspect of modern protein research. Element-specific detection at sub-ppb levels is essential to profile metal-binding proteins in complex matrices such as human serum. Coupling size exclusion chromatography (SEC) with inductively coupled plasma mass spectrometry (ICP-MS) provides simultaneous information on protein size and elemental composition, offering a powerful complement to conventional proteomics.

Goals and overview of the study

This collection of articles from the Agilent ICP-MS Journal Issue 27 (May 2006) presents multiple advances in ICP-MS technology and methodology:

• Development of an SEC-ICP-MS method for elemental profiling of pooled human serum according to protein mass fractions.
• Introduction of the EagleQuad quadrupole with an improved abundance sensitivity (AS) specification for the 7500 Series ICP-MS.
• Strategies to optimize sample throughput in ICP-MS by reducing wash-in/-out and data acquisition times.
• New features in Agilent ChemStation software Version B.03.03, notably Intelligent Rinse for automated background monitoring.
• Revised maintenance schedules for the 7500 Series to extend lens and octopole service intervals.
• Launch of the Agilent-branded ASX 520 autosampler with full Agilent support.

Methodology and instrumentation

SEC-ICP-MS analysis of pooled Type O human serum was performed on a Tosoh TSKgel 3000SWXL SEC column (5 μm, 25 nm pores) using 0.1 M sodium acetate buffer (pH 7.0) at 0.7 mL/min. A Peltier-cooled Agilent 1100 LC delivered samples (10 μL injections) monitored at 240 nm UV post-column. The LC effluent entered an Agilent 7500ce ICP-MS fitted with a GlassExpansion PFA OpalMist concentric nebulizer. Both LC and ICP-MS were controlled by Agilent ChemStation ICP-MS Top software.

Quadrupole improvements were realized by manufacturing hyperbolic rods (EagleQuad) to 1 μm tolerance, yielding AS specifications of 5×10⁻⁷ (Cs) at low mass and 1×10⁻⁷ (Cs) at high mass for the 7500 Series. Sample-introduction optimization included using reduced-ID peristaltic tubing (0.64 mm ID) and tailored rinse solutions (0.5% HCl for acidic rinse; ammonium hydroxide–EDTA–Triton X-100–H₂O₂ for alkaline rinse). Software enhancements in Version B.03.03 introduced Pre-emptive Rinse and Intelligent Rinse to minimize unnecessary rinse times by actively monitoring selected isotopes.

Main results and discussion

SEC-ICP-MS chromatograms of undiluted serum revealed three major metal-binding fractions: high-mass globular proteins (<83 kDa), the transferrin/HSA range (83–20 kDa), and low-mass peptides (>20 kDa). All nine monitored isotopes (including ⁵⁶Fe/⁵⁷Fe) were detected in at least one fraction. Sub-20 kDa peptides displayed three partially resolved sub-fractions, with Ni and Zn peaks approaching column resolution limits.

EagleQuad performance testing demonstrated record AS for quadrupole-based ICP-MS. Throughput optimization experiments indicated that reducing integration time from 0.3 to 0.1 s per isotope, selecting only essential isotopes, and minimizing stabilization delays can cut total run-to-run times by over an order of magnitude—for example, from 166 s down to 12.6 s for a 26-element panel.

ChemStation Version B.03.03’s Intelligent Rinse feature allowed dynamic termination of rinse cycles once user-defined background thresholds were reached, achieving faster sample cycles without compromising blank levels. Revised maintenance guidelines for the 7500ce extended lens cleaning intervals to 6 months and octopole checks annually under typical environmental workloads. The new Agilent ASX 520 autosampler offers fully supported hardware integration.

Benefits and practical applications

SEC-ICP-MS enables rapid, minimal-preparation profiling of metal-binding proteins in biofluids, facilitating biomarker discovery for diseases such as cancer and Alzheimer’s. Enhanced quadrupole AS improves trace-element detection in complex samples. Throughput improvements reduce per-sample analysis time and cost in high-volume laboratories. Intelligent Rinse and Pre-emptive Rinse capabilities increase uptime and consistency. Extended maintenance cycles and supported autosampler hardware lower operational overhead.

Future trends and possibilities

Advances in high-resolution chromatography and capillary SEC interfaces are expected to further resolve low-mass metal-peptide complexes. Integration with proteomic workflows combining structural elucidation (e.g., MS/MS) will deliver comprehensive metalloproteome maps. Continued development of reaction/collision cell chemistries and smart software automation will enhance interference removal and throughput. Fully networked autosampler systems and remote diagnostics may further streamline routine ICP-MS operations.

Conclusion

These developments demonstrate the versatility and evolving performance of ICP-MS for metallomic research and routine analysis. The combination of SEC separation, record-high abundance sensitivity, automated rinsing methods, software enhancements, and robust hardware support equips laboratories to conduct fast, sensitive, and reliable elemental profiling of biological and environmental samples.

Instrumentation used

• Agilent 1100 Series HPLC with Peltier cooling
• Tosoh TSKgel 3000SWXL SEC column (5 μm, 25 nm)
• Agilent 7500ce ICP-MS with EagleQuad quadrupole
• GlassExpansion PFA OpalMist concentric nebulizer
• Agilent ChemStation ICP-MS Top software Version B.03.03
• Agilent ASX 520 autosampler

References

1. Haraguchi H. J Anal Atom Spectrom 2003;19:5–14.
2. Wind M, Wolf DL. J Anal Atom Spectrom 2004;19:20–25.
3. Jakubowski N, Lobinski R, Moens L. J Anal Atom Spectrom 2004;19:1–4.
4. Lobinski R, Szpunar J. Anal Chim Acta 1999;400:3–21.
5. Dudev T, Lim C. Chem Rev 2003;103:773–787.
6. Coni E, Bocca B, Galoppi B, et al. Microchem J 2000;67:187–194.
7. Nischwitz V, Michalke B, Kettrup A. J Anal Atom Spectrom 2003;18:444–451.
8. Shiobara Y, Yoshida T, Suzuki KT. Toxicol Appl Pharmacol 1998;152:309–316.
9. Gallicchio L, Flaws JA, Sexton M, Ioffe OB. Toxicol Lett 2004;152:245–252.
10. Boulyga SF, Loreti V, Bettmer J, Heumann KG. Anal Bioanal Chem 2004;380:198–204.
11. Richarz A-N, Brätter P. Anal Bioanal Chem 2002;372:412–418.
12. Profrock D, Leonhard P, Ruck W, Prange A. Anal Bioanal Chem 2005;381:194–202.