A Multi-Well Method for the CD138 and AML/MDS FISH Testing of Multiple Biomarkers on a Single Slide in Multiple Myeloma and AML/MDS Patients

Background/Objectives: Genetic abnormalities play a pivotal role in patient risk stratification, therapeutic decision-making, and elucidating the disease pathogenesis in hematological malignancies. In multiple myeloma (MM) and acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS), numerous recurring genetic aberrations are well documented. Fluorescence in situ hybridization (FISH) is a cornerstone of clinical diagnostics for detecting these abnormalities. Conventionally, FISH assesses up to two biomarkers, with one or two circles per slide, but this approach faces challenges when cancer cell yields are limited, particularly in post-treatment follow-up specimens. Methods: To overcome this limitation, we developed a multi-well method, enabling the simultaneous testing of multiple biomarkers on a single microscopic slide. This study included 53 MM and 129 AML/MDS cases. Results: With a cohort of 182 patients, 1016 FISH assays performed on multi-well slides accurately detected diagnostic genetic aberrations previously identified by karyotyping and/or FISH, achieving a sensitivity and specificity of 100%. The use of multi-well slides achieved up to a 2.5-fold increase in the number of wells per slide while achieving more than a 3-fold reduction in the reagent volume compared to traditional methods. This advancement leverages distinct FISH signal patterns to strategically combine biomarkers within multiple wells, suitable for specimens from diagnosis, follow-ups, and relapses, regardless of the cancer cell quantity. Conclusions: The multi-well approach enhances the accessibility to comprehensive biomarker analysis, reducing both the processing time and costs. Beyond MM and AML/MDS, this technique holds promise for use with other hematological malignancies with limited sample volumes, offering an efficient, cost-effective solution for precision diagnostics.