Morphology and metaphase FISH assays on bone marrow of the variant APL. (A) Bone marrow morphology showing hypergranular promyelocytes lacking Auer rods. Image of May-Grünwald-Giemsa–stained smear was acquired using an Olympus BX-40 microscope (Olympus, Tokyo, Japan) equipped with a 100×/1.3 NA oil objective and a mounted DP12 digital camera (Olympus). (B) The LSI RARA dual-color break-apart rearrangement probe (Vysis, Downers Grove, IL) resulted in a normal fusion signal on one chromosome 17 and a small green signal close to the q terminal region of der(17) with deletion of the proximal red signal. (C) FISH with the LSI PML-RARA dual-color dual-fusion translocation probe (Vysis) resulted in normal red PML signals, one normal green RARA signal (not shown), and one split RARA signal. (D) A PRKAR1A probe (BAC RP11–120M18), labeled with Spectrum Orange (Vysis), combined with LSI PML-RARA dual-fusion translocation probe (as in panel C) confirmed that the fusion gene expressed by the patient was the result of a complex chromosome rearrangement which disrupted both the RARA gene on 17q21 and PRKAR1A on 17q24; the normal chromosome 17 has a single green signal (RARA) and a distal red signal (PRKAR1A), whereas the der(17) has a split green signal (RARA) where the more distal green signal colocalizes with a weaker red signal (PRKAR1A), indicating deletion of part of the probe. Note that the strong red PML probe signals on the 2 chromosomes 15 are because of image enhancement that was necessary to visualize the weak signals on the der(17). All FISH was performed according to manufacturer's instructions, and chromosomes were counterstained with DAPI (Sigma, St Louis, MO). Analysis was undertaken using a Zeiss Axioskop fluorescence microscope (Zeiss, Oberkochen, Germany) equipped with a 100× /1.3 NA oil objective. Images captured using the Isis image analysis system (MetaSystems, Altlussheim, Germany) were processed with FISH imaging system version 5.2 software (MetaSystems).