Abstract A032: Performance evaluation of a new protein stabilizing blood collection tube with a novel whole blood separation device for clinical plasma proteomics

Abstract Introduction: We have developed a novel blood separation device (BCD) which separates plasma from cellular components by lateral flow and stabilizes the sample for transport to a testing laboratory. Patient blood is drawn into an anticoagulant blood collection tube (BCT) and then applied to the separation device. There is outstanding interest in the field of clinical proteomics regarding the formulation in BCTs and whether they would also additionally the plasma proteome. This is especially critical where hemoglobin may provide interference to the analyte being measured. We evaluated the performance of a newly available plasma protein stabilizing BCT in combination with the BCD and other currently available K2EDTA BCT (no stabilization). Methods: Whole blood from normal healthy donors was drawn into protein stabilizing BCT and a comparator anticoagulant BCT. At baseline, 24-, 48-, 72- and 120-hour time points, a 250 μL aliquot of whole blood from each tube was spotted on to the BCD to allow separation of the cellular components and plasma. All of the test specimens on the spotted devices were allowed to dry overnight to replicate time in transit. After drying, proteins were subjected to MALDI-TOF MS (mass spectrometry) after extraction with water and co- crystallization with sinapinic acid. Spectra were acquired in triplicate and m/z feature intensities for hemoglobin were assessed in R. Results: Visual inspection of the new protein stabilizing and K2EDTA BCTs showed the steady accumulation of hemoglobin over time in the plasma layer in the latter tubes. After spotting on the BCD and MALDI-TOF analysis, plasma from the protein- stabilizing BCT showed no significant increases in either the hemoglobin α or β subunits over the five (5) day period. This was consistent across all donors (heme α p = 0.7933, heme β p = 0.6425) and at all timepoints assessed. In the K2EDTA BCT, both the α and β hemoglobin subunits increased significantly over time and across all donors (heme α p < 0.0001, heme β p = 0.0232). The hemoglobin α subunit accumulated in the plasma component to a greater degree than did the β subunit (heme α slope = 4.863E-5, heme β slope = 1.398E-5). These data suggest that the formulation in the protein stabilizing BCT may attenuate red blood cell hemolysis to a greater degree as compared to conventional K2EDTA blood collection tubes. Additionally, the mean reproducibility for both hemoglobin subunits was lower when the protein stabilizing BCT was used than when the anticoagulant BCT was used across all donors at all time points (protein stabilizing BCT x ̅ CV = 8.5%, anticoagulant BCT x ̅ CV = 8.9%). Conclusions: The use of the newly available protein stabilizing BCT with the BCD reduces the accumulation of hemoglobin in the plasma component while improving reproducibility when compared to the currently used anticoagulant BCT. Interference by heme was significantly attenuated as measured by a sensitive MALDI-ToF assay. Citation Format: Colin McDowell, Nylev Vargas, Gary A Pestano. Performance evaluation of a new protein stabilizing blood collection tube with a novel whole blood separation device for clinical plasma proteomics [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A032.