A practical criterion for focusing of unstained cell samples using a digital holographic microscope.

Digital holographic microscopy (DHM) is an important technique that may be used for quantitative phase imaging of unstained biological cell samples. Since the DHM technology is not commonly used in clinics or bioscience research labs, at present there is no well-accepted focusing criterion for unstained samples that users can follow while recording image plane digital holograms of cells. The usual sharpness metrics that are useful for auto-focusing of stained cells do not work well for unstained cells as there is no amplitude contrast. In this work, we report a practical method for estimating the best focus plane for unstained cells in the digital hologram domain. The method is based on an interesting observation that for the best focus plane the fringe pattern associated with individual unstained cells predominantly shows phase modulation effect in the form of bending of fringes and minimal amplitude modulation. This criterion when applied to unstained red blood cells shows that the central dip in the doughnut-like phase profile of cells is maximal in this plane. The proposed methodology is helpful for standardizing the usage of DHM technology across different users and application development efforts. LAY DESCRIPTION: Digital holographic microscopy (DHM) is slowly but steadily becoming an important microscopy modality and gaining acceptability for basic bio-science research as well as clinical usage. One of the important features of DHM is that it allows users to perform quantitative imaging of unstained transparent cells. Instead of using dyes or fluorescent labelling, DHM systems use quantitative phase as a contrast mechanism which depends on the natural refractive index variation within the cell samples. Since minimal wet lab processing is required in order to image cell samples with a DHM, cells can be imaged in their natural state. While DHM is gaining popularity among users, the imaging protocols across the labs or users need to be standardized in order to make sure that the same quantitative phase parameters are used for tasks such as quantitative phased based cell classification. One of the important operational tasks for any microscopy work is to focus the sample under study. While focusing comes naturally to users of brightfield microscopes based on image contrast, the focusing is not straightforward when samples are unstained so that they do not offer any amplitude contrast. When performing quantitative phase imaging, defocus can actually change the phase profile of the cell due to near-zone (Fresnel) diffraction effects. So unless a standardized focusing methodology is used, it will be difficult for multiple DHM users (potentially at different sites) to agree on quantitative results out of their phase images. DHM literature has prior works which perform numerical focusing of recovered complex wave-field in the hologram plane to find the best focus plane. However such methods are not user friendly and do not allow user the same focusing experience as in a brightfield microscope. The numerical focusing is therefore a reasonably good method for an optics researcher but not necessarily so for a microscopy technician looking at cell samples with a DHM system in a clinical setting. The present work provides a simple focusing criterion for unstained samples that works directly in the hologram domain. The technique is based on an interesting observation that the when an unstained cell sample is in the best-focus plane, its corresponding hologram (or fringe pattern) predominantly shows phase modulation manifested by bending of fringes at the location of the cell. This criterion can be converted into a simple numerical method as we have used to find the best-focus plane using a stack of through focus holograms. We believe that the technique can be used manually by visually observing the holograms or can be converted to an auto-focus algorithm for a motorized DHM system.

BIBI: Bayesian inference of breed composition.

The aim of this paper was to develop statistical models to estimate individual breed composition based on the previously proposed idea of regressing discrete random variables corresponding to counts of reference alleles of biallelic molecular markers located across the genome on the allele frequencies of each marker in the pure (base) breeds. Some of the existing regression-based methods do not guarantee that estimators of breed composition will lie in the appropriate parameter space, and none of them account for uncertainty about allele frequencies in the pure breeds, that is, uncertainty about the design matrix. To overcome these limitations, we proposed two Bayesian generalized linear models. For each individual, both models assume that the counts of the reference allele at each marker locus follow independent Binomial distributions, use the logit link and pose a Dirichlet prior over the vector of regression coefficients (which corresponds to breed composition). This prior guarantees that point estimators of breed composition such as the posterior mean pertain to the appropriate space. The difference between these models is that model termed BIBI does not account for uncertainty about the design matrix, while model termed BIBI2 accounts for such an uncertainty by assigning independent Beta priors to the entries of this matrix. We implemented these models in a data set from the University of Florida's multibreed Angus-Brahman population. Posterior means were used as point estimators of breed composition. In addition, the ordinary least squares estimator proposed by Kuehn et al. () (OLSK) was also computed. BIBI and BIBI2 estimated breed composition more accurately than OLSK, and BIBI2 had a 7.69% improvement in accuracy as compared to BIBI.

Gaussian covariance graph models accounting for correlated marker effects in genome-wide prediction.

Several statistical models used in genome-wide prediction assume uncorrelated marker allele substitution effects, but it is known that these effects may be correlated. In statistics, graphical models have been identified as a useful tool for covariance estimation in high-dimensional problems and it is an area that has recently experienced a great expansion. In Gaussian covariance graph models (GCovGM), the joint distribution of a set of random variables is assumed to be Gaussian and the pattern of zeros of the covariance matrix is encoded in terms of an undirected graph G. In this study, methods adapting the theory of GCovGM to genome-wide prediction were developed (Bayes GCov, Bayes GCov-KR and Bayes GCov-H). In simulated data sets, improvements in correlation between phenotypes and predicted breeding values and accuracies of predicted breeding values were found. Our models account for correlation of marker effects and permit to accommodate general structures as opposed to models proposed in previous studies, which consider spatial correlation only. In addition, they allow incorporation of biological information in the prediction process through its use when constructing graph G, and their extension to the multi-allelic loci case is straightforward.

Hypereosinophilic syndrome.

A case report, the clinical features, and a histopathological report in one case of Hypereosinophilic syndrome (HES) is reported. This patient had exfoliative dermatitis, urticaria and angiodema. She had received treatment with steroids which showed improvement and recurrence after stopping steroid therapy. Unique feature of this case was that skin eruptions were the main presentation. Skin eruptions may be the only manifestation of otherwise asymptomatic patient of HES.

Metabolites from apoptotic thymocytes inhibit thymopoiesis in adenosine deaminase-deficient fetal thymic organ cultures.

Murine fetal thymic organ culture was used to investigate the mechanism by which adenosine deaminase (ADA) deficiency causes T-cell immunodeficiency. C57BL/6 fetal thymuses treated with the specific ADA inhibitor 2'-deoxycoformycin exhibited features of the human disease, including accumulation of dATP and inhibition of S-adenosylhomocysteine hydrolase enzyme activity. Although T-cell receptor (TCR) Vbeta gene rearrangements and pre-TCR-alpha expression were normal in ADA-deficient cultures, the production of alphabeta TCR(+) thymocytes was inhibited by 95%, and differentiation was blocked beginning at the time of beta selection. In contrast, the production of gammadelta TCR(+) thymocytes was unaffected. Similar results were obtained using fetal thymuses from ADA gene-targeted mice. Differentiation and proliferation were preserved by the introduction of a bcl-2 transgene or disruption of the gene encoding apoptotic protease activating factor-1. The pan-caspase inhibitor carbobenzoxy-Val-Ala-Asp-fluoromethyl ketone also significantly lessened the effects of ADA deficiency and prevented the accumulation of dATP. Thus, ADA substrates accumulate and disrupt thymocyte development in ADA deficiency. These substrates derive from thymocytes that undergo apoptosis as a consequence of failing to pass developmental checkpoints, such as beta selection.

A study of electroencephalogram in meditators.

Electroencephalographic patterns were studied in 30 normal healthy individuals practicing meditation and compared with 10 normal healthy controls not practicing meditation. In this study, we found prominent alpha wave activity and increase it its voltage in meditators as compared to controls. Meditators had significantly more alpha rhythm as compared to control group. Percentage of alpha waves were higher in persons performing meditation with good coherence which suggested good homogenicity, uniformity and increased orderliness of brain.

CD73 expression and fyn-dependent signaling on murine lymphocytes.

CD73 is a glycosyl phosphatidylinositol-anchored protein with both ecto-enzyme activity (ecto-5'-nucleotidase) and signal transducing capabilities for human T lymphocytes. We now report an analysis of the distribution and function of CD73 in murine lymphoid tissues made possible by the development of the first monoclonal antibodies (mAb) specific for murine CD73. Subsets of T and B lymphocytes are CD73+ and the level of expression increases with lymphocyte maturation in both species. Among B cells, CD73 is largely restricted to cells which have undergone isotype switching. The signal transmitting function of CD73 is also conserved, as splenic T cells treated with anti-CD73 mAb plus phorbol 12-myristate 13-acetate proliferate and secrete IL-2. Fyn-/- mice are unresponsive to CD73 ligation, however, demonstrating the requirement for this tyrosine kinase in CD73-mediated signal transduction. CD73 is down-regulated after mAb plus cross-linking, suggesting that expression may be controlled by interaction with a ligand. Only small numbers of thymocytes are CD73+, so CD73 receptor functions are unlikely to be important for developing T cells. However, immunohistochemical analysis reveals that reticular and vascular cells throughout the thymus and other lymphoid tissues are markedly CD73+. Therefore, CD73 might mediate lymphocyte-stromal cell interactions or condition the local microenvironment to facilitate lymphocyte development and/or function.

ROLE OF EPIDURAL MEDICATION IN LUMBOSCIATIC SYNDROME.

Role of epidural medication through caudal route was studied in 109 patients having lumbago with or without sciatica to highlight the value of this mode of treatment which relieved symptoms in more than 70% of cases without hospitalisation and without being off work for long periods as in usual methods of conservative treatment.

Family planning and tuberculosis.

PIP: The mutual impact of India's family planning program and tuberculosis control program on each other was examined, and the conclusion reached that close cooperation between the 2 programs would have mutually beneficial results. The impact of the diagnosis of pulmonary tuberculosis on the family planning practices of 367 male and 290 female patients at a treatment clinic in Aligarh District was assessed. Prior to diagnosis only 10% of the patients practiced contraception, after diagnosis 19% practiced contraception, and after disease stabilization 29.5% practiced contraception. Among those patients who began practicing contraception after diagnosis, about 1/2 were motivated to practice by their physicians, and the remaining patients were motivated to practice either for economic reasons or because they feared spreading the infectious disease to an infant. About 2/3 of the tuberculosis patients had 3 or more children, and about 90% belonged to the lower socioeconomic class. Among female patients, disease onset occurred for 49.7% within 1 year of childbirth, and for 29.3%, it occurred during pregnancy. These findings suggested that an increase in the acceptance of family planning among members of the lower class and among high parity women would contribute toward decreasing the incidence of tuberculosis in the country. Clients who practiced contraception were also more likely to attend the tuberculosis treatment clinic on a more regular basis than non-practicing clients.^ieng

Silicosis in slate pencil workers. A clinicoradiologic study.

We present a study of 151 persons working in slate-pencil manufacturing industries located in the Mandsaur district of Madhya Pradesh, India. Cough, dyspnea, and pain in the chest were the important symptoms. Cyanosis, rhonchi, and crepitations were found in varying numbers of cases. The chest x-ray films were abnormal in 85 cases.