A major nucleolar protein B23 as a marker of proliferation activity of human peripheral lymphocytes.

A novel monoclonal antibody (Mab) (called 3C9) against a major nucleolar phosphoprotein B23 was used to study B23 qualitative and quantitative alterations in phytohemagglutinin (PHA) -stimulated human peripheral blood lymphocytes in indirect immunofluorescence and Western blots. It was shown that lymphocyte proliferation was accompanied by gradual augmentation of nucleoli and their accumulation of the protein B23 up to 2-fold by 16 h and 40-50 fold by 72 h, as compared with the non-stimulated cells. By parallel immunolabeling with the anti-Ki-67 antibody, it was shown that the early changes of B23 amount and localization occurred before an appearance of Ki-67 protein, a well-known marker of proliferating cells. Our results evidence that antibodies against B23 might be applied for recognition of human peripheral lymphocytes at early stages of their activation for proliferation, preceding the S-phase.

[Identification of nucleolar antigen in different types of cells using domestic monoclonal antibodies]. / Vyiavlenie iadryshkogo antigena v razlichnykh tipakh kletok s pomoshch'iu otechestvennykh monoklonal'nykh antitel.

Murine hybridoma 3C9 producing BCA-PC monoclonal antibodies and selectively staining the nucleoli in various cells, most intensely in actively proliferating cell cultures, was obtained by somatic hybridization of cells. BCA-PC MAb belong to IgM. As a rule, IgM antibodies are virtually unavailable for intracellular antigens, and therefore a special method has been developed for fixation and performance of indirect immunofluorescence with BCA-PC. By its molecular weight (37 and 43 kD) and distribution in the cellular nucleus, the antigen detected by BCA-PC resembles the nucleolar protein B23 located in the nucleolar granules. The content of B23 increases with malignant degeneration of cells and during cell proliferation. It is therefore believed that the new MAbs will help characterize the proliferative status of cells which determines the malignancy of a pathological process.

[Analysis of the proliferative activity of a cell using new monoclonal antibodies to nucleolar protein B23/nucleophosmin]. / Analiz proliferativnoi aktivnosti kletok s pomoshch'iu novykh monoklonal';nykh antitel k iadryshkovomu belku B23/nukleofozminu.

Nowadays, antinucleolar antibodies are widely used for exploration of the nucleolar organization and molecular mechanisms of ribosome production. Here we have described a new monoclonal antibody against the major nucleolar phosphoprotein B23/nucleophosmin (3C9) that is involved in the terminal stages of ribosome production. It is used to examine immunocytochemical peculiarities of the nucleolus in terms of the cell proliferative status and also during mitosis. In human peripheral blood lymphocytes, activated for proliferation with phytohaemagglutinin (PHA), PHA stimulation of lymphocytes was shown to result in accumulation of protein B23 in augmentative nucleoli. A comparative study of 3C9 and two other anti-B23 antibodies 20B2 and anti-B23 by Western blots and indirect immunofluorescence favored the idea that 3C9 cross-reacted with the major isoform of B23, B23.1, that have an apparent molecular weight of 40 kDa.

[Immunocytochemical method of assessment of the proliferative status of lymphoid cells in lymphoproliferative diseases]. / Immunotsitokhimicheskii metod otsenki proliferativnogo statusa limfoidnykh kletok pri limfoproliferativnykh zabolevaniiakh.

Search for the new approaches to the diagnosis of lymphoproliferative diseases and transition to the classification scheme "Working Classification of Non-Hodgkin's Lymphomas for Clinical Use" involved assessment of the proliferative status of pathological lymphoid cells by immunocytochemical method with the use of DAKO-PC (Ki-67) monoclonal antibodies. The course and specific features of immunocytochemical reaction with the use of peroxidase-antiperoxidase immune complexes and complexes of alkaline phosphatase and monoclonal antibodies to it on various types of preparations are described in detail, this appreciably extending the potentialities of the use of this method. The studies were carried out with lymphoid cells of different organs of the immune system and peripheral blood of 44 patients with lymphoproliferative diseases and of 5 donors. The results permit considering the developed and tried immunocytochemical method for assessment of the proliferative status of pathological cell populations informative as regards the use of the "Working Classification of Non-Hodgkin's Lymphomas".

[Evaluation of the proliferation and line of lymphoid cells in various lymphoproliferative diseases using a double immunocytochemical method]. / Otsenka proliferatsii i lineinoi prinadlezhnosti limfoidnykh kletok pri razlichnykh limfoproliferativnykh zabolevaniiakh dvoinym immunotsitokhimicheskim metodom.

The increased range of the diagnostic criteria and the introduction of the classification scheme "Practical Formulation of Non-Hodgkin's Lymphomas for Clinical Application" made the authors to study proliferative potential of lymphoid cells with identification of the cell line affiliation. The investigation was carried out with the use of the double immunocytochemical test employing anti-nuclear (Ki-67) and line-specific monoclonal antibodies on peripheral blood lymphocytes, those of lymph nodes and bone marrow obtained from 40 patients with lymphoproliferative diseases and on the cells of lymphoblastoid line 501. The disease variant was classified according to the WHO criteria. The highest (> 60%) proliferative activity of lymphoid cells was found in the bone marrow in acute non-T non-B lymphoblast leukemia and in lymph node in T-cell and non-T non-B-cell lymphoblast lymphosarcoma. A broad range in the number of proliferative cells existed among B-cell lymphosarcomas of lymphoblast (12-73%) and prolymphocytic (0-91%) variants. Together with heterogeneous immunological phenotype, this reflects a wide spectrum of various nosological entities recognized by the WHO classification as lymphoblast and prolymphocytic variants of lymphosarcoma, says about the defects of the above classification. Extremely low (< 1%) percent of proliferative cells was observed in B-cell chronic lymphoid leukemia and hairy-cell leukemia. These differences in tumor cell proliferative potential in different lymphoproliferative diseases agree with the views on maturation and differentiation of lymphoid cells. The data obtained by the authors support the necessity of correcting routine histological data by immunological characteristics of pathological lymphoid cells, primarily, by introduction of data on cell proliferative activity.(ABSTRACT TRUNCATED AT 250 WORDS)