Cytoplasmic NPM and multilineage involvement in NPMc⁺ AML. (Top, left) Myeloid sarcoma (skin, paraffin section). Nuclear plus aberrant cytoplasmic expression of NPM is seen in leukemic blasts. Normal residual vessel endothelial cells show the expected nucleus-restricted NPM expression (arrow). Immunostaining with monoclonal antibody 376 that recognizes both wild-type and mutated NPM (alkaline phosphatase anti–alkaline phosphatase [APAAP] technique; hematoxylin counterstaining; image was collected using an Olympus BX51 microscope and a UPlanApo 40×0.85 NA objective). (Top, right) NPMc⁺ AML (bone marrow biopsy, paraffin section). Cytoplasmic-restricted expression of mutant NPM protein in leukemic cells (arrow). A normal residual megakaryocyte is negative (double arrows). Immunostaining with a polyclonal antibody (Sil-A) that recognizes mutated but not wild-type NPM (APAAP technique; hematoxylin counterstaining; image was collected using an Olympus BX51 microscope and a UPlan FL 100×/1.3 NA oil objective). (Bottom, left) NPMc⁺ AML with multilineage involvement (bone marrow biopsy, image was collected using an Olympus BX51 microscope and a UplanApo 40×/0.85 NA objective). Double APAAP/immunoperoxidase staining for NPM (blue) and glycophorin (brown). Aberrant cytoplasmic expression of NPM is seen in proerythroblasts (arrow), megakaryocytes (double arrows), and myeloid blasts surrounding a blood vessel (asterisk). (Bottom, right) Higher magnification (image was collected using an Olympus BX51 microscope and a UPlan FL 100×/1.3 NA oil objective) from a different field of the same case. Proerythroblasts are strongly NPMc⁺ (blue) and weakly positive or negative for glycophorin (brown) (long arrow), while more mature erythroid precursors show nucleus-restricted NPM(blue)/strong surface glycophorin (brown) (short arrow). Double arrows indicate a NPMc⁺/glycophorin-negative megakaryocyte. A myeloid blast is positive for cytoplasmic NPM (blue) and negative for surface glycophorin (arrowhead).