Fast and easy: isolation of functional immune cells straight from mouse spleen
BORCZEWSKI A. 1, FLADE S. 1, DHANA E. 1, FODELIANAKI G. 1, EHRHARDT S. 1, WINKELS G. 1, BARANSKA A. 1
1 Miltenyi Biotec B.V. & Co. KG, Bergisch Gladbach, Germany
Backround: The isolation of lymphocytes and myeloid cells from the spleen of mice is a key technique in many basic, translational and applied studies. Research using magnetically isolated cells contributes to the understanding of the immune system, development of new vaccines and immunotherapeutic approaches. Therefore, choosing a reliable cell isolation method is crucial to ensure consistent and reproducible experimental results.
Objectives: Development of strategies that simplify and accelerate cell isolation workflows and ensure experimental reproducibility. These workflows should enable the isolation of highly viable and pure immune cells with good yields that retain their functionality.
Methods: Spleen dissociation and simultaneous magnetic labelling of cells (CD4, CD8, CD90.1, CD90.2, CD19, and CD11c) was performed using the GentleMACS dissociator. Subsequent automated cell isolation was done using autoMACS NEO Separator. To compare our innovative StraightFrom Spleen cell isolation with a competitor's untouched cell isolation approach, CD4 and CD8 T cell separation was performed in parallel. Target cell purity, yield, viability, cell activation status and functionality were assessed.
Results: CD4 and CD8 T-cell purity and viability were significantly higher for StraightFrom Spleen protocols compared to competitor protocols. The yield and viability of isolated cells was either superior for StraightFrom Spleen protocols or comparable for both protocols. No expression of CD25 or CD69 was observed when unstimulated CD4 and CD8 T cells were cultured overnight, and similar induction of these markers was observed after activation with the T cell Activation/Expansion Kit. Cells isolated using both protocols showed similar proliferation and expansion in cell culture.
Conclusion: We have developed a simple and fast cell isolation method with minimal hands-on time. Significantly accelerated isolation protocols enable rapid isolation of functional cells with excellent viability, purity and yield. The workflows presented here offer an attractive alternative to the laborious multi-step protocols commonly used in academic and industrial laboratories.