Production of a mouse monoclonal antibody reactive with a human nuclear antigen associated with cell proliferation
TL;DR: A first series of immunostainings of tumour biopsies indicated that Ki‐67 may be a potent tool for easy and quick evaluation of the proportion of proliferating cells in a tumour.
Abstract: The production of a mouse monoclonal antibody, Ki-67, is described. The Ki-67 antibody recognized a nuclear antigen present in proliferating cells, but absent in resting cells. Immunostainings with Ki-67 revealed nuclear reactivity in cells of germinal centres of cortical follicles, cortical thymocytes, neck cells of gastrointestinal mucosa, undifferentiated spermatogonia and cells of a number of human cell lines. The Ki-67 antibody did not react with cells known to be in a resting stage, such as lymphocytes, monocytes, parietal cells and Paneth's cells of gastrointestinal mucosa, hepatocytes, renal cells, mature sperm cells, brain cells, etc. Expression of the antigen recognized by Ki-67 could be induced in peripheral blood lymphocytes after stimulation with phytohaemagglutinin, whereas it disappeared from HL-60 cells stimulated with phorbol esters to differentiate into mature macrophages in a resting stage. These findings suggest that Ki-67 is directed against a nuclear antigen associated with cell proliferation. A first series of immunostainings of tumour biopsies indicated that Ki-67 may be a potent tool for easy and quick evaluation of the proportion of proliferating cells in a tumour.
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TL;DR: The Annexin V assay offers the possibility of detecting early phases of apoptosis before the loss of cell membrane integrity and permits measurements of the kinetics of apoptotic death in relation to the cell cycle.
5,291 citations
TL;DR: Although the Ki‐67 protein is well characterized on the molecular level and extensively used as a proliferation marker, the functional significance still remains unclear; there are indications, however, that Ki‐ 67 protein expression is an absolute requirement for progression through the cell‐division cycle.
Abstract: The expression of the human Ki-67 protein is strictly associated with cell proliferation. During interphase, the antigen can be exclusively detected within the nucleus, whereas in mitosis most of the protein is relocated to the surface of the chromosomes. The fact that the Ki-67 protein is present during all active phases of the cell cycle (G(1), S, G(2), and mitosis), but is absent from resting cells (G(0)), makes it an excellent marker for determining the so-called growth fraction of a given cell population. In the first part of this study, the term proliferation marker is discussed and examples of the applications of anti-Ki-67 protein antibodies in diagnostics of human tumors are given. The fraction of Ki-67-positive tumor cells (the Ki-67 labeling index) is often correlated with the clinical course of the disease. The best-studied examples in this context are carcinomas of the prostate and the breast. For these types of tumors, the prognostic value for survival and tumor recurrence has repeatedly been proven in uni- and multivariate analysis. The preparation of new monoclonal antibodies that react with the Ki-67 equivalent protein from rodents now extends the use of the Ki-67 protein as a proliferation marker to laboratory animals that are routinely used in basic research. The second part of this review focuses on the biology of the Ki-67 protein. Our current knowledge of the Ki-67 gene and protein structure, mRNA splicing, expression, and cellular localization during the cell-division cycle is summarized and discussed. Although the Ki-67 protein is well characterized on the molecular level and extensively used as a proliferation marker, the functional significance still remains unclear. There are indications, however, that Ki-67 protein expression is an absolute requirement for progression through the cell-division cycle.
4,359 citations
TL;DR: A cell atlas of mouse organogenesis provides a global view of developmental processes occurring during this critical period, including focused analyses of the apical ectodermal ridge, limb mesenchyme and skeletal muscle.
Abstract: Mammalian organogenesis is a remarkable process. Within a short timeframe, the cells of the three germ layers transform into an embryo that includes most of the major internal and external organs. Here we investigate the transcriptional dynamics of mouse organogenesis at single-cell resolution. Using single-cell combinatorial indexing, we profiled the transcriptomes of around 2 million cells derived from 61 embryos staged between 9.5 and 13.5 days of gestation, in a single experiment. The resulting ‘mouse organogenesis cell atlas’ (MOCA) provides a global view of developmental processes during this critical window. We use Monocle 3 to identify hundreds of cell types and 56 trajectories, many of which are detected only because of the depth of cellular coverage, and collectively define thousands of corresponding marker genes. We explore the dynamics of gene expression within cell types and trajectories over time, including focused analyses of the apical ectodermal ridge, limb mesenchyme and skeletal muscle. Data from single-cell combinatorial-indexing RNA-sequencing analysis of 2 million cells from mouse embryos between embryonic days 9.5 and 13.5 are compiled in a cell atlas of mouse organogenesis, which provides a global view of developmental processes occurring during this critical period.
1,865 citations
TL;DR: It is found that tumor-derived progenitors form neurospheres that can be passaged at clonal density and are able to self-renew, which may have important implications for treatment by means of specific targeting of stem-like cells within brain tumors.
Abstract: Pediatric brain tumors are significant causes of morbidity and mortality. It has been hypothesized that they derive from self-renewing multipotent neural stem cells. Here, we tested whether different pediatric brain tumors, including medulloblastomas and gliomas, contain cells with properties similar to neural stem cells. We find that tumor-derived progenitors form neurospheres that can be passaged at clonal density and are able to self-renew. Under conditions promoting differentiation, individual cells are multipotent, giving rise to both neurons and glia, in proportions that reflect the tumor of origin. Unlike normal neural stem cells, however, tumor-derived progenitors have an unusual capacity to proliferate and sometimes differentiate into abnormal cells with multiple differentiation markers. Gene expression analysis reveals that both whole tumors and tumor-derived neurospheres express many genes characteristic of neural and other stem cells, including CD133, Sox2, musashi-1, bmi-1, maternal embryonic leucine zipper kinase, and phosphoserine phosphatase, with variation from tumor to tumor. After grafting to neonatal rat brains, tumor-derived neurosphere cells migrate, produce neurons and glia, and continue to proliferate for more than 4 weeks. The results show that pediatric brain tumors contain neural stem-like cells with altered characteristics that may contribute to tumorigenesis. This finding may have important implications for treatment by means of specific targeting of stem-like cells within brain tumors.
1,810 citations
TL;DR: Results obtained indicate that Ki-1 antigen is an inducible lymphoid-associated molecule that identifies a group of hitherto poorly characterized normal and neoplastic large lymphoid cells in Hodgkin's disease and Disorders in which only a minority of cells express Ki- 1 antigen probably represent lesions in whichonly some of the abnormal cells have transformed into an "activation state.
1,779 citations
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TL;DR: The identification of an autoantibody in the sera of some patients with systemic lupus erythematosus that reacted with nuclear antigen(s) of proliferating cells that might serve as useful biologic markers to study stimulated lymphocytes and other proliferation cells.
Abstract: This study reports the identification of an autoantibody in the sera of some patients with systemic lupus erythematosus that reacted with nuclear antigen(s) of proliferating cells. The autoantibody was initially detected by the observation that it did not react in immunofluorescence with nuclei of renal tubular or glomerular cells, nor with hepatic parenchymal cells, but only reacted with scattered cells in the interstitial tissue of these two organs. In contrast, many lymphocytes in lymph node follicles, spleen, and thymus reacted with this antinuclear antibody. Normal peripheral blood lymphocytes did not show nuclear staining but after mitogenic stimulation, 20% of cells became positive. Nuclear staining was not restricted to lymphocytes but was also observed in several tissue culture cells lines such as Hep-2 cells (human epithelial carcinoma), Ehrlich ascites tumor cells, and baby hamster kidney cells. The reactive nuclear antigen(s) was soluble in physiologic saline and reacted with serum autoantibody to give a precipitin line in immunodiffusion that was immunologically distinct from DNA and other known nuclear antigen-antibody precipitating systems. Autoantibodies to proliferating cell nuclear antigen(s) might serve as useful biologic markers to study stimulated lymphocytes and other proliferating cells.
844 citations
TL;DR: Investigating the immunohistologic architecture of follicular centroblastic/centrocytic lymphoma showed a more or less close similarity to the organization of secondary follicles, which confirmed that this lymphoma was composed of a neoplastic T zone and a non-malignant B zone.
Abstract: Hoping to improve the systems for identifying and classifying normal and malignant lymphoid subpopulations, frozen and paraffin sections of nonmalignant lymphoid tissue and of malignant lymphomas were immunostained for surface (S) and cytoplasmic antigens using the peroxidase-antiperoxidase method. Primary follicle cells and follicle mantle cells known to be part of the recirculating B-cell pool were found to be constantly Ia and C3 receptor (C3R) positive, mostly SIgM and SIgD positive and cytoplasmic immunoglobulin (CIg) negative. The light zone of germinal centers (GC), which is rich in centrocytes, contained a large number of T cells and showed the well-known intercellular Ig network pattern; the dark zone, containing densely packed centroblasts, was usually free of T cells, but was bordered ay a mantle-like accumulation of T cells. Usually only some of the GC cells were definitely positive for SIg and CIg of different classes. All cells reacted positively for Ia and C3R. In areas described by other a...
388 citations
TL;DR: It is suggested that the hitherto unknown cell population detected with the monoclonal antibody Ki‐I in normal lymphoid tissue is the normal equivalent of H and SR cells.
Abstract: In this study the antigenic profile of Hodgkin (H) and Sternberg-Reed (SR) cells from cases of Hodgkin's disease was analysed using a large panel of monoclonal and polyclonal antibodies reactive with cells of lymphoid and haemotopoietic origin. The aim of this investigation was, firstly, to throw light on the origin of H and SR cells and, secondly, to determine whether there is any evidence to support recent suggestions that H and SR cells differ antigenically between different histological categories of Hodgkin's disease. Frozen sections (from 24 cases) and paraffin sections (83 cases) were stained by immunoenzymatic methods and the results compared with those obtained from staining a wide variety of reactive and neoplastic tissue samples (including examples of tuberculosis, sarcoidosis, malignant histiocytosis, histiocytosis X, osteomyelosclerosis and non-Hodgkin's lymphoma). The results revealed that H and SR cells of all types of Hodgkin's disease consistently lack markers found on null cells, B cells, T cells, cells of monocyte/macrophage series, interdigitating reticulum cells, dendritic reticulum cells and erythropoietic and thrombopoietic cells. However, H and SR cells constantly expressed an antigen detectable with the recently produced monoclonal antibody Ki-I. The vast majority of typical and lacunar type H and SR cells contained the granulocyte-related antigens detected by monoclonal antibodies TU5, TU6, TU9 and 3C4, whereas other more or less specific granulopoietic cell markers (such as peroxidase, chloroacetate esterade, lysozyme, cationic leukocyte antigen and OKMI) were consistently absent. H and SR cells in cases of nodular paragranuloma (nodular type of Hodgkin's disease with lymphocyte predominance) were not monotypic in light chain type (as has been previously reported), but rather contained chi and lambda chains within the same cells, as do typical and lacunar type H and SR cells. Immunostaining of normal and hyperplastic lymphoid tissue with the Ki-I antibody led to the detection of a new, as yet unidentified, small-cell population of unknown origin and function, which is present between, around, and within cortical follicles. It is concluded from these findings that H and SR cells constitute a unique cell type that differs in many properties from all other known cell types. Furthermore, H and SR cells of the various histological types of Hodgkin's disease are more closely related than previously believed. It is suggested that the hitherto unknown cell population detected with the monoclonal antibody Ki-I in normal lymphoid tissue is the normal equivalent of H and SR cells.
387 citations
TL;DR: The relationship of PCNA to proliferation and blast transformation may be associated with events related to DNA synthesis in these cells and the autoantibody was used as a reagent to determine the distribution.
Abstract: A nuclear antigen associated with cell proliferation (proliferating cell nuclear antigen-PCNA) and blast transformation is recognized by autoantibodies in the sera of some patients with systemic lupus erythematosus This autoantibody is a precipitating antibody and also reacts in immunofluorescence, staining the nucleoplasm of proliferating and blast-transformed cells The autoantibody was used as a reagent to determine the distribution of PCNA in a synchronized continuous B lymphoid cell line (WiL-2) and in mitogen-induced blast-transformed lymphocytes In WiL-2 cells, PCNA was detected as speckled nucleoplasmic staining in G1, S, and G2 phases of the cell cycle In addition, during late G1 and early S phases, PCNA was also detected in the nucleolus During mitogen-induced blast transformation of lymphocytes, PCNA was noticed in the nucleolus before the initiation of DNA synthesis and later became nucleoplasmic with disappearance of nucleolar staining These studies demonstrate that the relationship of PCNA to proliferation and blast transformation may be associated with events related to DNA synthesis in these cells
312 citations