RMD open
Synovial fluid cell isolation protocol affects immune cell recovery and composition across inflammatory states
By Tesfaye AM, Leta JT, Phung D and 8 others.
Abstract
Objective
Synovial fluid single-cell immunoprofiling is a powerful tool for investigating joint arthropathies. However, cell isolation protocols are yet to be standardised. We compared different cell-isolation protocols across diverse synovial fluid samples, evaluating cell recovery, sample inclusion rate and immune-cell composition.
Methods
From 48 synovial fluid samples with volumes>2 mL, we performed paired cell isolation of whole synovial fluid cells (WSFCs), with and without hyaluronidase and synovial fluid mononuclear cells (SFMCs) by density-gradient centrifugation. Of these, nineteen paired samples underwent deep immunophenotyping using a 31-marker spectral flow cytometry panel with dimensionality reduction and unsupervised clustering. Immune cell frequencies were quantified across protocols. Synovial fluid rheology was assessed in an additional 23 samples.