SJIP38: A single-tube 38-parameter spectral immunophenotyping panel for the lineage classification of pediatric leukemias

Flow cytometric immunophenotyping is central to the diagnosis and classification of pediatric leukemia but traditionally requires multiple tubes to evaluate the breadth of markers needed for lineage assignment and disease characterization. Here, we describe the development of a 38-parameter spectral flow cytometry immunophenotyping panel (SJIP38) for lineage classification of pediatric leukemias. The panel was designed for the initial evaluation of suspected de novo or relapsed acute leukemia in pediatric patients and consolidates markers previously distributed across a 13-tube conventional assay into a single high-dimensional spectral panel. The assay was designed to preserve established diagnostic markers, reduce sample requirements and tube number, and expand immunophenotypic resolution. Development was performed on the Sony ID7000 six-laser spectral cytometry platform using a systematic fluorochrome selection strategy guided by spillover spread characteristics, antigen density, and commercial availability of conjugated reagents. The resulting panel enables simultaneous assessment of lineage-defining, maturation-associated, and aberrantly expressed antigens across multiple pediatric leukemia subtypes. In addition to supporting routine diagnostic immunophenotyping, SJIP38 establishes a standardized fluorochrome palette and marker framework for future spectral assays, including measurable residual disease and expanded hematological malignancy panels.