The Impact of Detachment Methods (Trypsin, Accutase, and Accumax) on the Expression of Stem Cell Markers CXCR4 and CD146

Assaf, Wail Abou; Schmidt, Jan; Jouklová, Nela; Kapitán, Martin; Jankovičová, Karolína; Soukup, Tomáš

doi:10.14712/fb2025071030118

Fol. Biol. 2025, 71, 118-122

https://doi.org/10.14712/fb2025071030118

The Impact of Detachment Methods (Trypsin, Accutase, and Accumax) on the Expression of Stem Cell Markers CXCR4 and CD146

Wail Abou Assaf^1,2

, Jan Schmidt^1,2

, Nela Jouklová^1,2

, Martin Kapitán^1,2

, Karolína Jankovičová³

, Tomáš Soukup⁴

¹Department of Dentistry, Faculty of Medicine in Hradec Králové, Charles University, Hradec Králové, Czech Republic
²Department of Dentistry, University Hospital Hradec Králové, Hradec Králové, Czech Republic
³Department of Clinical Immunology and Allergology, University Hospital Hradec Králové, Hradec Králové, Czech Republic
⁴Department of Histology and Embryology, Faculty of Medicine in Hradec Králové, Charles University, Hradec Králové, Czech Republic

Received June 2025
Accepted July 2025

Trypsin (TRY) combined with ethylenediaminetetraacetic acid (EDTA) is a widely used dissociation agent due to its efficiency and cost-effectiveness. However, its impact on preserving stem cell marker expression, such as C-X-C chemokine receptor type 4 (CXCR4) (critical for cell migration and homing) and cluster of differentiation 146 (CD146) (involved in pluripotency and angiogenesis), may be suboptimal compared to alternatives such as Accutase (ACC) and Accumax (ACMX), as shown previously in bone marrow-derived stem cells (BM-MSCs). Limited data exist on these agents’ effects on dental pulp stem cells (DPSCs). This study aims to investigate the influence of TRY, ACC, and ACMX on the expression of CXCR4 and CD146 in DPSCs. Seven characterized DPSC lines were cultured under standardized conditions and detached using TRY-EDTA, ACC, or ACMX. The expression of CXCR4 and CD146 was quantified via multicolour flow cytometry using an innovative DURAClone SC panel with supplementary anti-CXCR4 antibody. Comprehensive statistical analyses were performed to evaluate differences in marker preservation. No statistically significant differences in CXCR4 or CD146 expression were observed across the detachment methods (P > 0.05). ACMX consistently demonstrated marginally higher mean expression levels for both markers (CXCR4: 84.77 %; CD146: 93.91 %) compared to ACC (CXCR4: 83.45 %; CD146: 93.41 %) and TRY (CXCR4: 83.95 %; CD146: 92.99 %). While differences were not statistically significant, ACMX consistently yielded higher mean expression of both CXCR4 and CD146, indicating a potential advantage in preserving marker integrity during the cell detachment.