Immunogold Labeling of Human Leukocytes for Scanning Electron Microscopy and Light Microscopy: Quantitative Aspects of the Methodology

When cell surface antigens are labeled with the colloidal gold marker, backscattered electron images (BEI) reveal all the gold particles and, therefore, permit total counts. Secondary electron images (SEI) show only a small percentage of the gold particles and are inadequate for quantitative evaluation. For determination of the cellular labelinq index, a time-consuming method implies the screening of 100 cells by scanninq electron microscopy, at a magnification of approximately 12,000 to 15,000x, with continuous SE/BE shifts. A much more efficient method is to transfer the SEM sample or its equivalent under the light microscope and to count the total number of gold labeled cells in the epi-polarization mode. The total cell count can be evaluated under UV light, taking advantage of the autofluorescence of the glutaraldehyde fixed cells

Immuno-Scanning Electron Microscopy of Normal and Leukemic Leukocytes Labeled with Colloidal Gold

The immunogold method, utilizing 40 nm colloidal gold particles which can be selectively visualized with the scanning electron microscope (SEM) in the backscattered electron imaging mode was used for the study of blood cells incubated with various monoclonal antibodies. Numerous antileukocyte monoclonal antibodies still recognize lightly glutaraldehyde prefixed antigens and can be used to identify various blood cell types and even to recognize their different maturation stages. Clearcut differences in surface morphology exist among peripheral blood normal leukocytes and even among the principal lymphocyte subclasses. Marked heterogeneity in surface morphology is, on the other hand, evident when studying precursors or leukemic cells. Immature cells show, nevertheless, relatively smooth surfaces while some distinct surface features appear on cells already committed toward a specific differentiation lineage. Hairy cells can also be precisely identified, especially when in small number in heterogeneous populations, combining their typical surface morphology with their positivity for B1 and Leu M5 monoclonal antibodies

Immuno-Cytochemistry with Backscattered Electrons

Some cytochemical reaction products are visible inside the cytoplasm of cells observed with the scanning electron microscope (SEM) using the backscattered electron imaging (BEI) mode. Methods can be utilized whenever they result in the deposition of heavy metal, like silver, lead or osmium at the sites of the enzymatic reaction. More recently the BEI mode of the SEM has been demonstrated to improve the detection of immunogold labeled cell surface antigens. Colloidal gold particles, 40 to 15 nm in diameter can be efficiently used for immuno-specific labeling. Moreover, cytochemical reactions can be applied to previously immunogold labeled cells, therefore combining the results of enzyme cytochemistry and of surface labeling at the level of each individual cell. The choice of fixative, incubation media, dehydration and drying methods should be guided by considerations on the sample characteristics for optimal electron scattering. Cytochemical as well as immuno-labeling reactions are not used per se but in combination with the study of cell surface morphology which needs, therefore, to be sufficiently well preserved. Coating should provide good conductivity and secondary electron emission, while emitting a minimal number of backscattered electrons. The application of these methods considerably enhances our capacity to characterize with the SEM the surface morphology of precisely identified subpopulations of many cell types

Testing the Properties of Radon Barrier Materials and Home Ventilation to Mitigate Indoor Radon

Indoor radon is the second cause of lung cancer. Mitigation strategies are based on (i) building protection with radon barrier materials, (ii) increasing home ventilation or (iii) room pressurization. A scale model room created with a porous ignimbrite rich in radon precursors was used as an analogue to test the indoor radon reduction ability of various radon barrier materials in a real room. The properties of these materials were tested with and without room pressurization by introducing outdoor air at different flow rates. The best materials reduced indoor radon up to 80% and, when the highest pressurization was applied, to 93%

Cell Surface Changes of Hemopoietic Cells During Normal and Leukemic Differentiation: An Immuno-Scanning Electron Microscopy Study

Hemopoietic cells display a wide range of cell surface antigens which are either lineage specific or acquired during differentiation. Monoclonal antibodies can be used, in conjunction with colloidal gold markers, to identify under the scanning electron microscopy (SEM) at the single cell level, specific lineage or maturation stages in the hemopoietic bone marrow. Normal bone marrow cells, either gradient separated or purified by immuno-magnetic methods and leukemic cell samples, which can be considered as frozen stages of hemopoietic differentiation, have been studied with this method. Typical cell surface morphologies, which characterize immature progenitor cells and cells committed or differentiated towards the lymphoid, myeloid, erythroid and megakaryocytic lineage have been identified. Correlations between cell surface features and some hemopoietic cells functions have been attempted on the basis of these findings

Scanning Electron Microscope Cytochemistry of Blood Cells

The backscattered electron imaging (BEI) mode of scanning electron microscopy (SEM) has been applied to study various histo-cytochemical reactions in biological specimens since the early seventies. Due to numerous, recent technical improvements the BEI mode of SEM now belongs to the routine of many SEM laboratories. For cytochemistry, BEI has been mainly used to: visualize intracellular structures and organelles; recognize the different cell types in heterogeneous populations or tissues; study the correlations between enzymatic activities and cell surface features. We have evaluated the most relevant results obtained in the study of blood cells and the possible future applications of these techniques

The impact of COVID-19 pandemic on pornography habits: a global analysis of Google Trends

As the COVID-19 spread globally, social distancing, self-isolation/quarantine, and national lockdowns have become crucial to control the pandemic. However, these measures may also lead to increases in social isolation, loneliness, and stress, which can alter the consumption of pornography habits. The aim of the study was thus to explore the interest pattern in pornography and coronavirus-themed pornography during the COVID-19 outbreak. Google Trends\uae was employed to determine the most popular porn websites (Porn, XNXX, PornHub, xVideos, and xHamster), and coronavirus-themed pornography worldwide and in six nations with different COVID-19 outbreak and self-isolation recommendations. We analyzed every search trend on Google\uae from January 9, 2020 to May 25, 2020 using \u201cjoint point regression analysis\u201d. Comparisons of week relative search volume (WRSV) and temporal patterns were analyzed to assess the change of interest in search terms during nations lockdowns. Paired t-test was used to compare WRSV values among the porn websites during the national lockdowns and the equivalent timespan of the weeks in the previous 4 years. The research trend of almost every keyword increased with significant inflection points for those nations with a straight \u201cstay at home orders\u201d (China, Italy, Spain, and France). \u201cPornHub\u201d and \u201cPorn\u201d showed the highest increase of interest worldwide with an average weekend percentage change (AWPC) of 4.9 and 3.8, respectively. The mean WRSV for keywords in USA and Sweden did not show a similar increase as the other nations. The WRSV percentage change with the historical data had a peak during the straight nations\u2019 lockdowns (p < 0.01). All the nations had a significant increase in WRSV coronavirus-themed pornography for each keyword (p < 0.01) with an AWPC, ranging worldwide between 18.5 and 61.8 (p < 0.01), after the beginning of self-quarantine. As strengths this study uses a big data technology to collect worldwide trend of interest, however, data are anonymous and do not allow analysis of subpopulation groups. In conclusion, we demonstrated an increased interest in pornography and coronavirus-themed pornography after the outbreak of COVID-19 in nations with a straight \u201cstay at home orders\u201d

Reduced intensity conditioning allogeneic transplant for advanced chronic lymphocytic leukemia

We report the preliminary results of 12 patients with advanced stage chronic lymphocytic leukemia (CLL) transplanted following reduced intensity conditioning (RIC. With a median of 22 months of follow-up, 9 patients are alive and 3 have died of progressive disease, graft-versus-host disease (GVHD) or toxic hepatitis. Acute grade I-III GVHD occurred in 33% of patients and chronic GVHD in 50%. Eight of the 12 patients achieved a complete remission (CR) and 2 patients a partial remission (PR). Donor lymphocyte infusion was effective in 6 patients. Event-free survival, progression-free survival and non-relapse mortality at 3 years were 68%, 42% and 16%, respectively. Our results show successful immunomodulation and reduction in tumor burden in high risk CL

Induction of neurotrophin expression via human adult mesenchymal stem cells: implication for cell therapy in neurodegenerative diseases.

In animal models of neurological disorders for cerebral ischemia, Parkinson's disease, and spinal cord lesions, transplantation of mesenchymal stem cells (MSCs) has been reported to improve functional outcome. Three mechanisms have been suggested for the effects of the MSCs: transdifferentiation of the grafted cells with replacement of degenerating neural cells, cell fusion, and neuroprotection of the dying cells. Here we demonstrate that a restricted number of cells with differentiated astroglial features can be obtained from human adult MSCs (hMSCs) both in vitro using different induction protocols and in vivo after transplantation into the developing mouse brain. We then examined the in vitro differentiation capacity of the hMSCs in coculture with slices of neonatal brain cortex. In this condition the hMSCs did not show any neuronal transdifferentiation but expressed neurotrophin low-affinity (NGFRp75) and high-affinity (trkC) receptors and released nerve growth factor (NGF) and neurotrophin-3 (NT-3). The same neurotrophin's expression was demonstrated 45 days after the intracerebral transplantation of hMSCs into nude mice with surviving astroglial cells. These data further confirm the limited capability of adult hMSC to differentiate into neurons whereas they differentiated in astroglial cells. Moreover, the secretion of neurotrophic factors combined with activation of the specific receptors of transplanted hMSCs demonstrated an alternative mechanism for neuroprotection of degenerating neurons. hMSCs are further defined in their transplantation potential for treating neurological disorders

‘Verdello’, ‘Verdicchio’ and ‘Verduschia’: an example of integrated multidisciplinary study to clarify grapevine cultivar identity

‘Verdello’, ‘Verdicchio’ and ‘Verduschia’ are registered in the official Italian Catalogue as three distinct grapevine varieties. Twenty-five accessions of these cultivars, encompassing known or presumed synonyms, coming from CRA repositories and from vineyards where they are traditionally cultivated, have been genotyped with eleven SSR markers. For morphological comparison, one accession for each variety has been described with 57 characters of OIV 2009 list; phenological and yield traits have also been recorded. In addition, the phenotypic comparison has been extended to the literature descriptions. The same DNA profile has been obtained for all 25 accessions; moreover, present and historical ampelographic data showed a very high similarity. All this information leads to the conclusion that these three varieties are, in fact, the same cultivar