Comparison of two peripheral mononuclear cell isolation protocols for flow cytometry crossmatching

Area of Science:

• Immunology
• Transplantation Science
• Clinical Diagnostics

Background:

• Flow cytometry crossmatching is crucial in organ transplantation to assess antibody-mediated rejection risk.
• Mononuclear cell separation is a key step in some crossmatching protocols, with Ficoll-Hypaque being a standard but time-consuming method.
• Faster methods for cell separation are needed to improve laboratory efficiency in clinical settings.

Purpose of the Study:

• To compare the efficacy of the SepMate tube method versus the standard Ficoll-Hypaque protocol for mononuclear cell isolation in flow cytometry crossmatching.
• To evaluate the impact of sample storage duration (0, 24, 48, 72 hours at room temperature) on the performance of both separation methods.

Main Methods:

• Samples from organ transplant donors were collected and stored at room temperature for varying durations.
• Mononuclear cells were isolated using either the SepMate tube or the Ficoll-Hypaque protocol.
• Isolated cells were used for flow cytometry crossmatching, with and without pronase treatment.

Main Results:

• The SepMate tube method demonstrated reliable performance for flow cytometry crossmatching when samples were stored at room temperature for 48 hours or less.
• The Ficoll-Hypaque protocol remained effective for samples stored at room temperature for more than 48 hours.
• Both methods showed comparable results within their respective optimal storage timeframes.

Conclusions:

• The SepMate tube offers a faster and efficient alternative to Ficoll-Hypaque for mononuclear cell isolation in flow cytometry crossmatching for samples stored up to 48 hours.
• The choice of method should be guided by sample storage duration to ensure accurate crossmatching results in clinical organ transplantation.
• Optimizing cell separation techniques can enhance the workflow and turnaround time for critical pre-transplant assessments.