Development of a kit for urine collection on filter paper as an alternative for Pompe disease screening and monitoring by LC-HRMS

Anal Methods. 2023 Aug 17;15(32):3932-3939. doi: 10.1039/d3ay00587a.

PMID: 37539791

Hygor M R de Souza, Fernanda B Scalco, Rafael Garrett

Highlights: This study investigates a newly developed filter paper based-urine collection kit to facilitate specimen shipment, and liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) analysis to determine Glc4 and creatinine in dried urine on filter paper.

Abstract

Background: A lack of the α-glucosidase acid enzyme results in Pompe disease (PD), an inborn metabolic error. It has a severe negative influence on the health and quality of life of patients and has the potential to cause infant death. Urinary glucose tetrasaccharide (Glc4) screening by high performance-liquid chromatography is one of the well-established diagnostic techniques that has been useful in monitoring Glc4 levels in patients receiving enzyme replacement treatment and demonstrating the efficacy of therapy. However, the specimen shipping process from a sample collecting location to a specialized laboratory for monitoring the Glc4 is costly and presents preanalytical challenges.

Methods: For the sake of specimen shipping, we created a kit for collecting urine on filter paper, and we used liquid chromatography and high-resolution mass spectrometry (LC-HRMS) analysis to find the amounts of Glc4 and creatinine in dried urine on filter paper. The LC-HRMS was successfully designed and validated on the basis of combining targeted and untargeted screening on the same specimen injection. There was a strong correlation between Glc4 and creatinine in dried and liquid urine samples, according to Bland-Altman statistics. Glc4 and other metabolites in dried urine showed stability for at least 7 days at 4 and 22 °C, and 3 days at 50 °C. By sending the kit by post, it was put to the test under real-world conditions to see how stable the analytes were and how effective it was.

Conclusion: The stability of the compounds was maintained after two days of transit without refrigeration, demonstrating the accuracy of the urine collection kit and method of analysis to identify the PD biomarker Glc4.

Keywords: Pompe disease, screening, biomarker, urine collection