Quantitation of infliximab using clonotypic peptides and selective reaction monitoring by LC-MS/MS

Int Immunopharmacol. 2015 Sep;28(1):513-20. doi: 10.1016/j.intimp.2015.07.007. Epub 2015 Jul 25.


Objectives: Although therapeutic concentrations of infliximab (chimeric IgG1 kappa) are associated with improved clinical prognosis, clinical laboratory methods for monitoring are limited. Therefore, we aimed to develop a LC-MS/MS method to measure infliximab in serum.

Methods: Infliximab was measured using isotope-labeled peptides and horse IgG as a surrogate internal standard. After trypsin digestion, peptides were separated by reverse-phase C8 LC and detected by MS/MS on an ABSciex API 5000; analyte-to-internal standard peak area ratios were used for quantitation. Sera from patients receiving infliximab were collected at different time points in treatment and compared with a commercially available ECLIA method.

Results: The linear dynamic range of the assay was 1-100 μg/mL (R(2)>0.998); both intra- and inter-assay imprecisions were <20%. Patients undergoing infliximab therapy had trough concentrations of 8.5 ± 8.8 μg/mL (mean ± SD), which substantially increased 48-72 h after infusion (77 ± 40 μg/mL), then fell after 28-32 days (15 ± 11 μg/mL). A comparison of LC-MS/MS and ECLIA methods demonstrated a slope of 0.967 (95% CI: 0.894-1.034, r=0.970).

Conclusions: We have demonstrated the ability to quantify infliximab in patients using clonotypic peptides. This approach has the potential to be quickly adaptable to other monoclonal antibodies and to expand the availability of testing for this class of therapeutics in routine clinical practice.

Keywords: Infliximab; Mass spectrometry; Monoclonal antibody therapeutics; Selective reaction monitoring.

MeSH terms

  • Anti-Inflammatory Agents / blood*
  • Antirheumatic Agents / blood*
  • Chromatography, Liquid
  • Humans
  • Immunoglobulin G
  • Infliximab / blood*
  • Peptides
  • Tandem Mass Spectrometry


  • Anti-Inflammatory Agents
  • Antirheumatic Agents
  • Immunoglobulin G
  • Peptides
  • Infliximab