The Elephant in the Analysis: Post-Acquisition Matrix Adjustment and the Transparency Gap in High-Parameter Cytometry

High-parameter and spectral flow cytometry increase sensitivity to errors in compensation and spectral unmixing. Foundational guidance has emphasized that matrices should be generated to ensure measurement accuracy rather than adjusted based on visual appearance. However, recent community discussions acknowledge that limited post-acquisition correction may be appropriate when transparently reported and its analytical impact documented. This study systematically evaluated reporting practices for post-acquisition compensation or spectral unmixing adjustment across all Optimized Multicolor Immunophenotyping Panels (OMIPs) published in Cytometry Part A (OMIP-001 to OMIP-119). The MIFlowCyt Author Checklist and associated supplementary materials were reviewed for explicit evidence of post-calculation matrix modification. Post-acquisition adjustment was reported in 23 of 119 OMIPs (~ 19%) across both conventional and spectral platforms. Adjustments were most commonly guided by NxN inspection, fluorescence-minus-one controls, and expected biological expression patterns. Reporting detail varied widely and was frequently qualitative. These findings demonstrate variability in current reporting practices and support the need for consistent documentation of matrix evaluation and any post-acquisition modification.