Fast and accurate quantitation of intact IGF-1 from serum by HRAM LC-MS for clinical research

Importance of the Topic

Insulin-like growth factor 1 (IGF-1) is a critical polypeptide hormone involved in human growth regulation, pediatric endocrinology diagnostics and anti-doping testing. Accurate measurement of intact IGF-1 in serum is essential for assessing growth hormone disorders, monitoring therapeutic interventions and detecting illicit performance-enhancing use.

Objectives and Overview

This study evaluates a high-resolution accurate-mass (HRAM) workflow combining the Thermo Scientific Vanquish UHPLC platform with the Orbitrap Exploris 120 mass spectrometer. The goal is to demonstrate fast, robust and precise quantitation of intact IGF-1 in human serum for clinical research applications.

Methodology

• Sample Preparation: Addition of ^15N-labeled IGF-1 internal standard, protein precipitation with acidic ethanol, cryo-precipitation after Tris neutralization. Calibration standards prepared in PBS/20% acetonitrile/1% BSA.
• Chromatographic Separation: Automated online SPE heart-cutting; Hypersil GOLD C8 analytical column (50 × 2.1 mm, 5 µm); 3 min total run time; gradient 80:20 to 5:95 (A: 0.1% formic acid in water, B: 0.1% formic acid in acetonitrile); flow 0.6 mL/min; column 35 °C; injection 50 µL.
• Mass Spectrometry Detection: Orbitrap Exploris 120 in SIM mode monitoring the 7+ charge state of IGF-1 (m/z 1093.5225) and ^15N-IGF-1 internal standard (m/z 1106.7671); resolution 60 000 (FWHM); EASY-IC source; isolation window 2 m/z; mass extraction width 5 ppm.

Used Instrumentation

• Thermo Scientific Vanquish Flex UHPLC system
• Thermo Scientific Orbitrap Exploris 120 mass spectrometer with EASY-IC ion source
• Thermo Scientific Hypersil GOLD C8 column (50 × 2.1 mm, 5 µm)
• MilliporeSigma Chromolith RP-18e trap cartridge

Main Results and Discussion

The method achieved a lower limit of quantitation (LLOQ) of 10 ng/mL with linearity over 15.6–2 000 ng/mL (r² > 0.995). Over 54 batches and 130 hours of continuous operation, calibration bias remained within ±6%, precision (CV) < 7%, and mass accuracy stable within 1.5 ppm for calibrators and 2 ppm for spiked serum samples. No significant carryover or matrix interference was observed, demonstrating excellent system robustness and reproducibility.

Benefits and Practical Applications of the Method

• High-throughput 3-minute analysis suitable for large clinical cohorts
• Enhanced selectivity and sensitivity for intact IGF-1 in complex matrices
• Long-term stability over hundreds of injections without maintenance
• Applicability in clinical diagnostics, pediatric endocrinology studies and anti-doping laboratories

Future Trends and Potential Applications

Further improvements in HRAM MS speed and sensitivity along with automated sample preparation will enable multiplexed profiling of peptide hormones. Integration of advanced data processing algorithms and machine learning could expand IGF-1 quantitation into personalized medicine, comprehensive endocrine panels and real-time anti-doping surveillance.

Conclusion

The combination of Vanquish UHPLC and Orbitrap Exploris 120 MS provides a fast, accurate and robust workflow for intact IGF-1 quantitation in serum, fulfilling clinical research and anti-doping requirements with exceptional performance over extended sequences.

References

No external references provided.