A Targeted LC-MS/MS-Based Quantitative Assay for Detecting Plasma Factor XIII A/B Subunit Deficiency.

Area of Science:

• Biochemistry
• Hematology
• Analytical Chemistry

Background:

• Factor XIII (FXIII) is essential for stabilizing blood clots through fibrin crosslinking.
• Deficiencies in FXIII subunits (FXIIIA or FXIIIB) can cause bleeding disorders.
• Current assays primarily measure only the FXIIIA subunit, limiting comprehensive FXIII assessment.

Purpose of the Study:

• To develop and validate a targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for quantifying both FXIIIA and FXIIIB subunits in plasma.
• To assess the clinical utility of the developed LC-MS/MS assay for diagnosing FXIII deficiency.

Main Methods:

• Development and validation of a targeted LC-MS/MS assay for FXIIIA and FXIIIB quantification.
• Selection of peptides based on analytical robustness, sensitivity, linearity, and precision.
• Comparison with a conventional ammonia release assay and analysis of 98 clinical plasma samples.

Main Results:

• The LC-MS/MS assay demonstrated high sensitivity (LOD: 1.56–3.67 mU/mL), linearity (r² > 0.998), and precision (CV <10%).
• Strong correlation (r = 0.983) and agreement (Cohen κ = 0.846) were observed when compared to the conventional FXIIIA assay.
• The assay successfully identified FXIII deficiency cases, showing reduced FXIIIA and variable FXIIIB levels in clinical samples.

Conclusions:

• The validated LC-MS/MS assay provides accurate quantification of both FXIIIA and FXIIIB subunits.
• This assay enhances diagnostic accuracy for Factor XIII deficiency.
• The method offers potential for further research into FXIII dynamics in various pathological conditions.