LC-MS/MS Quantification of Intact Insulin-Like Growth Factor I (IGF-I) in Serum for Clinical Research

Importance of the Topic

The accurate measurement of insulin-like growth factor I is critical in clinical research and endocrine studies where IGF-I serves as a key biomarker for growth hormone activity and metabolic regulation. Reliable quantification in serum or plasma informs diagnosis of growth disorders, monitoring of therapy, and population studies.

Study Objectives and Overview

This work aims to develop a robust, digestion-free LC-MS/MS method for direct quantification of intact IGF-I in serum. The approach seeks to simplify sample preparation, enhance sensitivity and specificity, and achieve a broad dynamic range suitable for clinical research.

Methodology

Sample pretreatment involves denaturation of binding proteins using sodium dodecyl sulfate followed by protein precipitation with acidified acetonitrile. A mixed-mode solid-phase extraction on a 96-well µElution plate enables cleanup without affinity steps. Chromatographic separation employs a sub-2-µm solid-core C18+ column with a rapid aqueous–organic gradient. Tandem quadrupole detection in positive electrospray mode uses optimized multiple reaction monitoring transitions of the 7+ and 8+ charge states of IGF-I to ensure selectivity.

Applied Instrumentation

• 96-well Oasis MAX µElution solid-phase extraction plate
• ACQUITY UPLC I-Class chromatography system
• CORTECS UPLC C18+ 2.1×50 mm, 1.6 µm column
• Xevo TQ-XS tandem quadrupole mass spectrometer with ESI+

Main Results and Discussion

The optimized workflow achieved over 90 percent recovery of intact IGF-I with a lower limit of quantification of 5 ng/mL in mouse plasma and consistent performance in human serum. Calibration curves were linear (r2>0.99) across 5–1,000 ng/mL (mouse) and 100–1,000 ng/mL (human). Intra-day precision was below 7 percent RSD and accuracy within 94–108 percent. The use of the CORTECS C18+ column halved peak widths (<6 seconds) and quadrupled signal-to-noise compared to a traditional fully porous C18 phase.

Benefits and Practical Applications

• Rapid sample preparation under 1.5 hours without enzymatic digestion or affinity capture
• High sensitivity and selectivity for endogenous and elevated IGF-I levels
• Compatibility with high-throughput workflows and standard LC-MS platforms
• Cost-effective alternative to immunoassays with reduced reagent consumption

Future Trends and Potential Applications

Advances may include multiplexed analysis of related growth factors and binding proteins, automation of the µElution SPE workflow, and integration with high-resolution mass spectrometry for structural characterization. Continued development of charged-surface and core-shell columns promises further gains in throughput and sensitivity.

Conclusion

The presented LC-MS/MS assay offers a streamlined, reproducible, and sensitive solution for direct quantification of intact IGF-I in serum. By eliminating digestion and affinity steps, it accelerates assay development, reduces costs, and supports robust biobehavioral and clinical research applications.

References

1. Lopes F, Cowan DA, Thevis M, Thomas A, Parkin MC. Quantification of Intact Human Insulin-Like Growth Factor-I in Serum by Nano-UHPLC/MS/MS. Rapid Commun Mass Spectrom. 2014;28:1426–1432.
2. Bidlingmaier M, Friedrich N, Emeny RT, et al. Reference Intervals for Insulin-Like Growth Factor-1 From Birth to Senescence using an Automated Chemiluminescence Immunoassay. J Clin Endocrinol Metab. 2014;99(5):1712–1721.