Confocal microscopy

Chapter focusing on confocal microscopy. A confocal microscope is one in which the illumination is confined to a small volume in the specimen, the detection is confined to the same volume and the image is built up by scanning this volume over the specimen, either by moving the beam of light over the specimen or by displacing the specimen relative to a stationary beam. The chief advantage of this type of microscope is that it gives a greatly enhanced discrimination of depth relative to conventional microscopes. Commercial systems appeared in the 1980s and, despite their high cost, the world market for them is probably between 500 and 1000 instruments per annum, mainly because of their use in biomedical research in conjunction with fluorescent labelling methods. There are many books and review articles on this subject ( e.g. Pawley ( 2006) , Matsumoto( 2002), Wilson (1990) ). The purpose of this chapter is to provide an introduction to optical and engineering aspects that may be o f interest to biomedical users of confocal microscopy

Generalized valence bond wave functions in quantum Monte Carlo

We present a technique for using quantum Monte Carlo (QMC) to obtain high quality energy differences. We use generalized valence bond (GVB) wave functions, for an intuitive approach to capturing the important sources of static correlation, without needing to optimize the orbitals with QMC. Using our modifications to Walker branching and Jastrows, we can then reliably use diffusion quantum Monte Carlo to add in all the dynamic correlation. This simple approach is easily accurate to within a few tenths of a kcal/mol for a variety of problems, which we demonstrate for the adiabatic singlet-triplet splitting in methylene, the vertical and adiabatic singlet-triplet splitting in ethylene, 2+2 cycloaddition, and Be_2 bond breaking

An enigmatic satellite

McLaughlin and Chadwick describe an X-linked tandem repeat that is transcribed, conserved and defies X inactivation. Is it selfish, functional or just an oddity

Correlated and geographically predictable Neanderthal and Denisovan legacies are difficult to reconcile with a simple model based on inter-breeding.

Although the presence of archaic hominin legacies in humans is taken for granted, little attention has been given as to how the data fit with how humans colonized the world. Here, I show that Neanderthal and Denisovan legacies are strongly correlated and that inferred legacy size, like heterozygosity, exhibits a strong correlation with distance from Africa. Simulations confirm that, once created, legacy size is extremely stable: it may reduce through admixture with lower legacy populations but cannot increase significantly through neutral drift. Consequently, populations carrying the highest legacies are likely to be those whose ancestors inter-bred most with archaics. However, the populations with the highest legacies are globally scattered and are unified, not by having origins within the known Neanderthal range, but instead by living in locations that lie furthest from Africa. Furthermore, the Simons Genome Diversity Project data reveal two distinct correlations between Neanderthal and Denisovan legacies, one that starts in North Africa and increases west to east across Eurasia and into some parts of Oceania, and a second, much steeper trend that starts in Africa, peaking with the San and Ju/'hoansi and which, if extrapolated, predicts the large inferred legacies of both archaics found in Oceania/Australia. Similar 'double' trends are observed for the introgression statistic f 4 in a second large dataset published by Qin and Stoneking (Qin & Stoneking 2015 Mol. Biol. Evol. 32, 2665-2674 (doi:10.1093/molbev/msv141)). These trends appear at odds with simple models of how introgression occurred though more complicated patterns of introgression could potentially generate better fits. Moreover, substituting archaic genomes with those of great apes yields similar but biologically impossible signals of introgression, suggesting that the signals these metrics capture arise within humans and are largely independent of the test group. Interestingly, the data do appear to fit a speculative model in which the loss of diversity that occurred when humans moved further from Africa created a gradient in heterozygosity that in turn progressively reduced mutation rate such that populations furthest from Africa have diverged less from our common ancestor and hence from the archaics. In this light, the two distinct trends could be interpreted in terms of two 'out of Africa' events, an early one ending in Oceania and Australia and a later one that colonized Eurasia and the Americas

Standing-wave-excited multiplanar fluorescence in a laser scanning microscope reveals 3D information on red blood cells

Standing-wave excitation of fluorescence is highly desirable in optical microscopy because it improves the axial resolution. We demonstrate here that multiplanar excitation of fluorescence by a standing wave can be produced in a single-spot laser scanning microscope by placing a plane reflector close to the specimen. We report that the relative intensities in each plane of excitation depend on the Stokes shift of the fluorochrome. We show by the use of dyes specific for the cell membrane how standing-wave excitation can be exploited to generate precise contour maps of the surface membrane of red blood cells, with an axial resolution of ~90 nm. The method, which requires only the addition of a plane mirror to an existing confocal laser scanning microscope, may well prove useful in studying diseases which involve the red cell membrane, such as malaria.Comment: 15 pages, 4 figures; changed the discussion of narrow-band detected fringes (Fig. 3) to describe the phenomenon as a moire pattern between the excitation and emission standing-wave fields, rather than a beats pattern; added DiI(5)-labelled red blood cell in Fig. 4 to show that standing-wave fringes are present even when the dye excitation wavelength is outside the haemoglobin absorption ban

Electron microscopic measurement of the size of the optical focus in laser scanning microscopy

We describe a method for measuring the lateral focal spot size of a multiphoton laser scanning microscope (LSM) with unprecedented accuracy. A specimen consisting of an aluminum film deposited on a glass coverslip was brought into focus in a LSM and the laser intensity was then increased enough to perform nanoablation of the metal film. This process leaves a permanent trace of the raster path usually taken by the beam during the acquisition of an optical image. A scanning electron microscope (SEM) was then used to determine the nanoablated line width to high accuracy, from which the lateral spot size and hence resolution of the LSM can be determined. To demonstrate our method, we performed analysis of a multiphoton LSM at various infrared wavelengths, and we report measurements of optical lateral spot size with an accuracy of 20 nm, limited only by the resolution of the SEM

Label Noise: Correcting a Correction

Training neural network classifiers on datasets with label noise poses a risk of overfitting them to the noisy labels. To address this issue, researchers have explored alternative loss functions that aim to be more robust. However, many of these alternatives are heuristic in nature and still vulnerable to overfitting or underfitting. In this work, we propose a more direct approach to tackling overfitting caused by label noise. We observe that the presence of label noise implies a lower bound on the noisy generalised risk. Building upon this observation, we propose imposing a lower bound on the empirical risk during training to mitigate overfitting. Our main contribution is providing theoretical results that yield explicit, easily computable bounds on the minimum achievable noisy risk for different loss functions. We empirically demonstrate that using these bounds significantly enhances robustness in various settings, with virtually no additional computational cost

ANALISIS SOSIAL KOMUNITAS ONLINE KOMSEL JUNIOR GKKA-I BALIKPAPAN

Pandemi Covid-19 yang melanda di dunia tentunya membawa berbagai dampak negatif kepada kehidupan masyarakat. Banyak aktivitas masyarakat yang terganggu karena adanya pandemi Covid-19 seperti dalam pekerjaan, kesehatan, ekonomi, dan masih banyak lagi lainnya. Salah satu aktivitas masyarakat yang terkena dampak Covid-19 adalah Komsel Junior GKKA-I Balikpapan yang aktivitasnya menjadi terganggu. Adanya perkembangan teknologi yang pesat pada saat ini membuat Komsel Junior GKKA-I Balikpapan menggunakan media Zoom sebagai alternatif untuk mengatasi pandemi Covid-19 yang membuat pertemuan secara offline tidak bisa dijalankan. Namun seiring berjalannya waktu Komsel Junior juga berjalan setengah offline supaya jalannya pertemuan lebih efektif. Analisis ini menggunakan pendekatan Participation Action Research. Hasil dari analisis ini terbagi menjadi beberapa analisis yaitu analisis geografi, analisis demografi, sejarah, struktur, mata pencaharian, stakeholder, masalah, dan tujuan. yang bertujuan untuk memahami komunitas ini secara lebih baik

Effectiveness of Non-Monetary Motivations on Job Performance in Tanzania: A Case of Institutions in Tabora Municipality, Tanzania

This study sought to examine “the effectiveness of non-monetary motivations on job performance in public institutions; a case of Tabora Municipal Council. The study was confined to four specific objectives which were: (1) To examine the process of Recognizing workers as non-monetary motivation on job performance in Tabora Municipal Council (2) To examine the process of Promoting workers as non-monetary motivation on job performance in Tabora Municipal Council (3) To determine the process of Training workers as non-monetary motivation on job performance in Tabora Municipal Council (4) To determine the relationship between non-monetary motivations and employees’ job performance in Tabora Municipal Council. A descriptive correlational study design was adopted and a total sample of 148 respondents was consulted. The stratified and purposive sampling techniques were used to select respondents. Questionnaires were employed as tools of data collection. Quantitative data were analyzed by frequencies, mean, standard deviation, and Pearson correlation, and the data were presented in tables and figures. The study findings revealed that the processes of non-monetary motivations follow labor and employment policy guidelines at Tabora Municipality. Furthermore, the study findings revealed that factors for successful practices are the use of fairness, policy, and evaluation feedback reports. The study findings revealed that there is a positive relationship between non-monetary motivations and job performance. Lastly, the study recommends that Trust from all employees must be taken into account to create a fair and equal chance when it comes to the awarding process. Management members should be trained properly to conduct the employee performance review as one of the tool requirements in granting non-monetary motivation awards to workers.&nbsp

Signatures of Introgression across the Allele Frequency Spectrum.

The detection of introgression from genomic data is transforming our view of species and the origins of adaptive variation. Among the most widely used approaches to detect introgression is the so-called ABBA-BABA test or D-statistic, which identifies excess allele sharing between nonsister taxa. Part of the appeal of D is its simplicity, but this also limits its informativeness, particularly about the timing and direction of introgression. Here we present a simple extension, D frequency spectrum or DFS, in which D is partitioned according to the frequencies of derived alleles. We use simulations over a large parameter space to show how DFS carries information about various factors. In particular, recent introgression reliably leads to a peak in DFS among low-frequency derived alleles, whereas violation of model assumptions can lead to a lack of signal at low frequencies. We also reanalyze published empirical data from six different animal and plant taxa, and interpret the results in the light of our simulations, showing how DFS provides novel insights. We currently see DFS as a descriptive tool that will augment both simple and sophisticated tests for introgression, but in the future it may be usefully incorporated into probabilistic inference frameworks