Optimized isolation of renal plasma cells for flow cytometric analysis

Area of Science:

• Immunology
• Cell Biology

Background:

• Plasma cells (PCs) are crucial for antibody production and immunity but also implicated in autoimmune diseases.
• PCs are typically found in bone marrow and spleen, but can infiltrate inflamed tissues like kidneys in conditions such as systemic lupus erythematosus (SLE).
• Accurate analysis of PCs in inflamed organs requires reliable single-cell suspension preparation methods.

Purpose of the Study:

• To evaluate the impact of enzymatic digestion on plasma cell surface marker integrity.
• To establish an optimized protocol for preparing viable renal single-cell suspensions for PC analysis.
• To compare the efficacy of mechanical disruption versus enzymatic digestion for immune cell isolation from kidneys.

Main Methods:

• Comparison of enzymatic digestion (collagenase) with mechanical disruption for isolating renal immune cells.
• Assessment of plasma cell surface marker expression (CD138, CD267/TACI) using flow cytometry.
• Evaluation of cell yield and purity for different cell populations (lymphocytes, stromal cells).

Main Results:

• Enzymatic digestion, particularly with collagenase, led to the loss of critical plasma cell surface markers (CD138, CD267/TACI).
• Mechanical tissue disruption provided a method for preparing viable renal single-cell suspensions without compromising PC surface marker integrity.
• Mechanical preparation enhanced the enrichment of lymphocytic immune cells, including plasma cells, for accurate flow cytometry and cell sorting.

Conclusions:

• Enzymatic digestion is detrimental to the reliable characterization of renal plasma cells due to surface marker loss.
• Mechanical disruption is a preferred method for preparing renal single-cell suspensions for accurate flow cytometry and analysis of viable plasma cells.
• The choice of tissue preparation method significantly impacts the yield and characterization of specific immune cell populations from solid organs.