Abstract
Background:
There is increasing interest in better understanding the immune cell composition of human milk and how these cells interact with neonatal immune development. However, consistent methods for immune cell isolation from human milk are lacking.
Research aim:
Our aim was to systematically compare available cell isolation techniques to isolate T lymphocytes from human milk samples.
Methods:
This repeated measures study design compared three cell isolation methods using human milk samples: (1) centrifugation, (2) immunomagnetic bead isolation, and (3) a combination of both methods. We assessed the proportion and viability of CD3+, CD4+, CD25+ and regulatory T cells using flow cytometry in isolated cells to compare the performance of these isolation methods.
Results:
Immunomagnetic separation is a feasible method to isolate T lymphocytes in human milk, similar to blood. It improves target cell enrichment and cell viability compared to centrifugation, which may be an advantage when the goal is to characterize rare cell types or when cells are further used in functional assays. No excess cell activation (CD25 positivity) was observed with the use of magnetic beads.
Conclusion:
Immunomagnetic separation of human milk T lymphocytes may have advantages over centrifugation depending on the intended downstream use of cells.