Delayed Processing of Secretin-Induced Pancreas Fluid Influences the Quality and Integrity of Proteins and Nucleic Acids

• Type of Material: Article
• Authors: Zobedia Cruz-Monserrate, Ohio State University; Kristyn Gumpper, Ohio State University; Sabrina Kaul, Ohio State University; Niharika Badi, Ohio State University; Samantha Terhorst, Ohio State University; Kelly Dubay, Ohio State University; Gregory Lesinski, Emory University; William Fisher, Baylor College of Medicine; Amy McElhany, Baylor College of Medicine; Luis F Lara, Ohio State University; Somashekar Krishna, Ohio State University; Thomas Mace, Ohio State University; Natalia Higuita-Castro, Ohio State University; Lilibeth Ortega-Pineda, Ohio State University; Michael A Freitas, Ohio State University; Alice Hinton, Ohio State University; Dhiraj Yadav, University of Pittsburgh; Phil A Hart, Ohio State University; Stephen J Pandol, Cedars-Sinai Medical Center; Saima Ahmed, Harvard Medical School; Benoit Fatou, Harvard Medical School; Hanno Steen, Harvard Medical School; Darwin L Conwell, Ohio State University
• Language: English
• Date: 2021-01-01
• Publisher: LIPPINCOTT WILLIAMS & WILKINS
• Publication Version: Author Accepted Manuscript (After Peer Review)
• Copyright Statement: © 2020 Wolters Kluwer Health, Inc. All rights reserved.
• License: Creative Commons Attribution-NonCommercial 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1097/MPA.0000000000001717
• Title of Journal or Parent Work: PANCREAS
• Volume: 50
• Issue: 1
• Start Page: 17
• End Page: 28
• Grant/Funding Information: This work was also supported in part by the Pelotonia Fellowship Program (Kaul and KD), by grant P30 CA016058 NCI and by the ChiRhoClin Research Institute, Inc. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Pancreas Foundation, the National Institutes of Health, the Pelotonia Fellowship Program or the ChiRhoClin Research Institute.; Research in this publication was supported by: The National Pancreas Foundation (ZC-M), The National Cancer Institute (NCI) R01CA223204 (ZC-M), and by the NCI and National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) under award number U01DK108327 (DC, ZC-M, PH, LL, Krishna, HS), U01DK108306 (DY).
• Keywords: IDENTIFICATION; Science & Technology; Life Sciences & Biomedicine; SYNTHETIC PORCINE SECRETIN; TANDEM; EARLY-DIAGNOSIS; JUICE SAMPLES; Gastroenterology & Hepatology; sample degradation; CONSORTIUM; BIOMARKERS; pancreatic cancer; ASSOCIATION; CANCER; biomarkers; pancreatitis; pancreatic fluid; COLLECTION

Abstract

Objectives Endoscopic pancreatic function tests are used to diagnose pancreatic diseases and are a viable source for the discovery of biomarkers to better characterize pancreatic disorders. However, pancreatic fluid (PF) contains active enzymes that degrade biomolecules. Therefore, we tested how preservation methods and time to storage influence the integrity and quality of proteins and nucleic acids. Methods We obtained PF from 9 subjects who underwent an endoscopic pancreatic function test. Samples were snap frozen at the time of collection; after 1, 2, and 4 hours on ice; or after storage overnight at 4°C with or without RNase or protease inhibitors (PIs). Electrophoresis and mass spectrometry analysis determined protein abundance and quality, whereas nucleic acid integrity values determined DNA and RNA degradation. Results Protein degradation increased after 4 hours on ice and DNA degradation after 2 hours on ice. Adding PIs delayed degradation. RNA was significantly degraded under all conditions compared with the snap frozen samples. Isolated RNA from PF-derived exosomes exhibited similar poor quality as RNA isolated from matched PF samples. Conclusions Adding PIs immediately after collecting PF and processing the fluid within 4 hours of collection maintains the protein and nucleic acid integrity for use in downstream molecular analyses.