Ultrastructural cytochemistry as a tool for studying transcriptional mechanisms

Ultrastructural cytochemistry is a powerful tool for investigating the biology of the cell nucleus. Thanks to its very high resolution, it has been possible to localize with extreme accuracy the sites of transcription, splicing and maturation of both mRNA and rRNA, their precise location as well as their movements. By means of immunocytochemical techniques, many nuclear proteins have been given a specific role in the transcriptional mechanisms, with the possibility of precisely mapping their location on a single RNA fibril. Starting from the 70s, the techniques have evolved from the resolution of autoradiography to that of gold-coupled antibodies, reaching a resolution of few nanometers. The use of correlative microscopy techniques as well as of electron tomography has also allowed the 3D imaging of ribonucleoprotein-containing structures in situ, in the nucleus. The association of a biological mechanism with the cytochemical localization of a specific molecule has been crucial in defining the functional organization of the cell nucleus, and has been invaluable for the understanding of many biological processes.

Use of Transmission Electron Microscopy in Diagnosis of Acute Leukemias: A Prospective Study of Fifty Cases.

Ultrastructural studies have contributed substantially to the understanding of the cellular morphology of the acute leukemias. We present here 50 cases (aged 3-55 years, M: F-32: 18) of acute leukemias, which were studied for morphology, conventional cyto chemistry, immunophenotyping, and transmission electron microscopy (TEM) including ultrastructural cytochemistry using myeloperoxidase (MPO) and platelet peroxidase activity. TEM morphology using ultrastructural cytochemistry of MPO helped diagnose three cases of acute myeloid leukemia with minimal myeloid differentiation (AML M0). The blasts showed large round nuclei, 1–2 nucleoli, chromatin with peripheral condensation and abundant mitochondria. Of total of 5 cases of acute promyelocytic leukemias (APML), all had strong Sudan Black, MPO, dual esterase positivity; one case was non-specific esterase positive and sensitive to fluoride. On TEM, this unusual case was identified to be microgranular variant of APML. TEM morphology and ultrastructural cytochemistry using platelet peroxidase helped diagnose 3 cases of AML M7 (acute megakaryocytic leukemia), 2 cases of acute biphenotypic leukemias and also in differentiating one case of acute proerythroblastic leukemia (AML M6b) from 3 cases of AML M6a or acute erythroleukemia. Thus, TEM is helpful in differentiating further the subgroups of AML-M5 and AML-M6, in identifying the microgranular variant of APML, and in confirming the diagnosis of AML-M0 and biphenotypic leukemia. Also, in cases with very hypercellular marrow and with associated myelofibrosis, where the bone marrow aspiration gives low cell count, TEM and ultrastructural cytochemistry are a valuable aid to arriving at a accurate diagnosis. Characteristics of acute leukemia cases (n=50) FAB Subtype Morphology, Conventional Cytochemistry and Immunophenotyping Transmission Electron Microscopy (?) = doubt in diagnosis AML-M0 3 (?) 3 AML-M1and M2 11 11 AML-M3 4, 1(?) 5: Hypergranular-4, hypogranular-1 AML-M4 5 5 AML-M5 5 5a-4, 5b-1 AML-M6 4 6a-3 (erythroleukemia), 6b-1 AML-M7 3 3 Biphenotypic 2 (?) 2 ALL-L1 8 8 ALL-L2 4 4