Determination of CD26 marker on human lymphocytes, previously treated with cytokinesTrpin, Ela (Avtor) Božič, Borut (Mentor) Varela-Calviño, Rubén (Komentor) Introduction: Dipeptidyl peptidase IV (DPPIV), also known as CD26, is a multifunctional membrane glycoprotein, expressed on cell surface of various cells, including T lymphocytes. Its surface expression on T lymphocytes is closely related to cell activation and differentiation. There’s an up-regulation of the marker on Th17 and Th1 cells and a down-regulation on Th2 cells. Research aim: The aim of this study was to determine DPPIV surface and intracellular expression on different helper T subsets (Th0, Th1, Th2, Th17) and find a correlation between them. Our objective was also to determine the amount of enzyme released to the medium during the incubation and the necessity of CD4+ isolation step. Methods: PBMCs for our experiments were obtained form 9 healthy individuals. For the determination of CD26 expression on different Th subtypes, cells were stained with antibodies against CD3, CD4, CD45RO and CD26 for surface analysis and anti-CD26 antibodies for intracellular analysis. Analysis of DPPIV expression on lymphocytes was done using flow cytometry. The presence of the enzyme in the cultivation medium after a 3-day cultivation was determined with ELISA and its activity with enzyme activity test. Results and discussion: The highest CD26 surface expression was observed on Th17 population, followed by Th1, Th0, and Th2 subtypes, with a significant difference between Th17 and Th2 subsets (p<0.05). The highest CD26 intracellular expression was observed on Th1 subtype, followed by Th0, Th2, and Th17 with no significant differences between them. The isolation step was much more important for intracellular analysis, where cells were stained only with antibodies against CD26, compared to surface analysis, where our target population was selected based on the presence of other membrane markers. The presence and the activity of the enzyme in the cultivation medium after a 3-day incubation were not high enough to be detected by any of the methods in most of our samples. Conclusion: Although we did not manage to find a correlation between CD26 surface and intracellular expression on different helper T subsets, we solved some other questions, among them the importance of CD4+ isolation step for intracellular analysis. It was evident that at the time of maximum surface expression of CD26, its leaking to the medium was still negligible.[E. Trpin]20182020-09-18 12:47:05Magistrsko delo/naloga120363UDK: 577.152.34+616-097(043.3)COBISS_ID: 4656497sl