Imaging Flow Cytometric Assessment of del(17p) and amp(1q21) in Circulating Plasma Cells of Myeloma Patients.

Multiple myeloma is a bone marrow-derived neoplastic proliferation of plasma cells with low levels of circulating tumor cells. Cytogenetic abnormalities are present in >90% of patients, when assessed by fluorescence in situ hybridization (FISH) on bone marrow, and the specific abnormalities provide vital prognostic information. Provided is a protocol for assessing the high-risk abnormalities del(17p) and amp(1q21) in the circulating plasma cells of myeloma patients by flow cytometry. This utilizes positive plasma cell identification by standard immunophenotyping and then chromosomal analysis by FISH using an imaging flow cytometer. Integrating cell phenotype and FISH in one test, a method called "immuno-flowFISH" allows for the detection of cytogenetic abnormalities in plasma cells identified by their antigenic profile. This method can be applied to both bone marrow and blood samples to detect primary abnormalities [i.e., hyperdiploidy; immunoglobulin heavy locus (IGH) translocation] and secondary abnormalities [i.e., del(17p) found in 10% of patients, and gain(1q) present in ∼40% of patients with myeloma]. Here we describe the protocol for the simultaneous detection of these secondary abnormalities in blood and bone marrow samples from myeloma patients that enables detection of single or "double hit" abnormalities, with the latter classified as ultra high-risk disease. The protocol includes the data analysis strategy, as well as the statistics used to confirm the presence of these abnormalities. Applying this method will facilitate blood-based monitoring for the presence and evolution of these critical cytogenetic abnormalities. © 2026 Wiley Periodicals LLC. Basic Protocol: Immuno-flowFISH assessment of del(17p) and amp(1q21) in multiple myeloma Support Protocol: Validation of single fluorophore for positive markers in multiple myeloma.