T and B cells are essential components of the adaptive immune system and are pivotal in both disease research and the development of new therapies. However, the current method of isolating these cells from whole blood involves density gradient centrifugation, which is time-consuming and operator-dependent, leading to inconsistent results.
To overcome these challenges, we devised the innovative REAlease technology protocol that allows for direct isolation of cells enabling the removal of superparamagnetic beads leading to highly pure unlabeled leukocytes. By integration of the new autoMACS NEO Separator into the workflow we could increase standardization of the cell separation and reduce hands on time to a minimum.
Positive isolation of T and B cells directly from human whole blood was performed using CD3, CD8 or CD19 StraightFrom Whole Blood REAlease MicroBead Kit. Purity and yield of cells as well as accessibility of the previously labeled epitope were determined. Viability, activation status and functionality of the isolated cells were assessed using in vitro assays.
With CD3, CD8 and CD19 StraightFrom Whole Blood REAlease MicroBead Kit T and B cells could be enriched with purities greater than 95% also from larger blood samples or even complete buffy coats. Analysis of activation markers CD25 and CD69 for T cells, CD80, CD86 and MHCII for B cells showed that enriched, unstimulated cells were not activated by the procedure. Further functional assays show full downstream functionality of isolated cells.
The REAlease isolation strategy combined the strengths of positive enrichment with the benefits of negative isolation methods, minimizing cell manipulation and reducing downstream process interference from remaining labeling.
In conclusion, our easy, fast, reliable, and automatable workflow provides a promising alternative for T and B cell isolation. This protocol could contribute significantly to the advancement of disease research and the development of novel therapies.