Aim: Protein quantitation by digestion of a biological sample followed by LC–MS analysis of a signature peptide can be a challenge because of the high complexity of the digested matrix. Results/methodology: The use of LC with high-resolution (quadrupole-TOF) MS detection allowed quantitation of the 22-kDa biopharmaceutical somatropin in 60 μl of rat plasma down to 25 ng/ml with minimal further sample treatment. Reducing the mass extraction window to 0.01 Da considerably decreased the interference of tryptic peptides, enhanced sensitivity and improved accuracy and precision. Analysis with LC–MS/MS resulted in a less favorable limit of quantitation of 100 ng/ml. Conclusion: HRMS is an interesting option for the quantitation of proteins after digestion and has the potential to improve sensitivity with minimal method development.
Keywords:
- digestion
- high-resolution mass spectrometry
- HRMS
- mass extraction window
- MEW
- rhGH
- somatropin