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Fluorescence-activated cell sorting (FACS) is a branch of flow cytometry that allows for the isolation of specific cell populations that can then be further analyzed by single-cell RNA sequencing (scRNA-seq). When utilizing FACS for population isolation prior to sequencing, it is essential to consider the protection of RNA from RNase activity, environmental conditions, and the sorting efficiency to ensure optimum sample quality. This study aimed to optimize a previously published MDSC flow cytometry strategy to FACS sort canine Myeloid-Derived Suppressor Cells (MDSC) with various permutations of RNAlater ™ and RiboLock™ before and after FACS sorting. Concentrations of RNAlater™ greater than 2 % applied before flow analysis affected cell survival and fluorescence, whereas concentrations ≤ 2 % and time ≤ 4 h had little to no effect on cells. To shorten the procedural time and to enhance the sorting of rare populations, we used a primary PE-conjugated CD11b antibody and magnetic column. The combination of RiboLock™ pre- and post-sorting for FACS provided the best quality RNA as determined by the RNA integrity number (RIN ≥ 7) for scRNA-seq in a normal and dog and a dog with untreated oral melanoma dog. As proof of principle, we sequenced two samples, one from a normal dog another from a dog with untreated oral melanoma. Applying scRNA-Seq analysis using the 10X Genomic platform, we identified 6 clusters in the Seurat paired analysis of MDSC sorted samples. Two clusters, with the majority of the cells coming from the melanoma sample, had genes that were upregulated (> log2); these included MMP9, MMP1, HPGD, CPA3, and GATA3 and CYBB, CSTB, COX2, ATP6, and COX 17 for cluster 5 and 6 respectively. All genes have known associations with MDSCs. Further characterization using pathway analysis tools was not attempted due to the lower number of cells sequenced in the normal sample. The benefit deriving from the results of the study helped to gain data consistency when working with cells prone to RNase activity, and the scRNA-seq provided data showing transcriptional heterogeneity in MDSC populations and potentially identifying previously unreported or rare cell populations.

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Marine mammals evoke strong public affection as well as considerable scientific interest. However, the resultant close contact with marine wildlife poses human health risks, including traumatic injury and zoonotic disease transmission. The majority of zoonotic marine mammal diseases result in localized skin infections in man that resolve spontaneously or with appropriate medical therapy. However, other marine mammal zoonoses, if left untreated, induce life-threatening systemic diseases that could pose public health risks. As the number of zoonotic diseases rises, the diagnosis of and treatment for these emerging pathogens pose special challenges requiring the expertise of physicians, veterinarians and wildlife biologists. Here, we provide a comprehensive review of the bacterial, viral and fungal marine mammal zoonotic diseases that we hope will be utilized by public health professionals, physicians, veterinarians and wildlife biologists to better understand, diagnose and prevent marine mammal zoonotic diseases.

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The measurement of the mobility of SF-6 in the mixtures SF6 -Ar and SF6 -Xe is reported over the density-reduced electric field strength E/N 1-180 Td (1 Townsend = 10(-17) V cm(2)), from a time-resolved pulsed Townsend technique. Simultaneously, the mobility of SF-6 in the same binary mixtures has been calculated from a set of collision cross sections for SF-6 -Ar, SF-6 -Xe, and SF-6 - SF6 using a Monte Carlo simulation procedure for ion transport. The good agreement between measured and calculated mobilities in these gas mixtures has led us to conclude that the validation of our cross section sets is confirmed. The elastic collision cross section, a predominant process for ion energies lower than about 10 eV, was determined from a semiclassical JWKB approximation using a rigid core potential model for the ion-neutral systems under consideration. This elastic cross section was then added to several other inelastic collision cross sections found in the literature for ion conversion, electron detachment of SF-6 and charge transfer. Moreover, the calculations of the mobility and the ratios of the transverse and longitudinal diffusion coefficients to the mobility were extended into a much wider E/N range from 1 to 4000 Td. Additionally, we have also calculated the energy distribution functions and the reaction coefficients for ion conversion and electron detachment. Finally, we have shown that the range of validity for the calculation of the mobility in gas mixtures from Blanc's law is only valid for the low E/N region, where the interaction is dominated by elastic collisions and the ion distribution function remains essentially Maxwellian.

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The principle of microscopic time reversal of physical processes, detailed balance, is widely used and depended upon in the theoretical community as a connection between photorecombination (PR) and photoionization (PI). This paper reports on a novel use of detailed balance and the comparison of experimental results obtained by two very distinct techniques to determine both the metastable fraction of an ion beam and partial Sc3+ PR cross sections and partial Sc2+ PI cross sections for the ground state and for two metastable states. The Sc2+ to Sc3+ system presents a unique opportunity to obtain comprehensive state-selective information by combining results from PR and PI experiments.

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High-resolution absolute experimental measurements and two independent theoretical calculations were performed for photoionization of O+ ions from the 2P(o) and 2D(o) metastable levels and from the 4S(o) ground state in the photon energy range 30-35.5 eV. This is believed to be the first comparison of experiment and theory to be reported for photoionization from metastable states of ions. While there is correspondence between the predicted and measured positions and relative strengths of the resonances, the cross-section magnitudes and fine structure are sensitive to the choice of basis states.

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The purpose of this study was to describe the dental caries experience in 12-year-old students in the State of Mexico and to detect changes after 9 years of salt fluoridation (1988-1997). The 1987 WHO dental caries criteria were used in both surveys. The population studied encompassed 2,275 12-year-old students in 1988, and 1,138 in 1997. The proportion of 'caries-free' children was 10. 3% in the first survey and 27.7% in the second one. The mean DMFT index was 4.39 (SD 2.9) in 1988 and 2.47 (SD 2.4) in 1997, the confidence interval of the differences between DMFT means was [95% CI 1.73, 2.11]; the caries reduction detected was 43.7%. The Unmet Restorative Index was 82.3% in 1988 and 72.8% in 1997. The results of the present study indicate that the oral health status of State of Mexico students has improved during the last decade; however, there is still a need for further caries reduction and an increment in access to dental treatment.

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EEG spectral parameters were computed in a group of children with different degrees of difficulty in learning to read and write. For statistical analyses, Z-transformed values according to normative age-regression equations were used to control the age effects. Canonical Correlation Analysis between absolute power in different bands and the categories of the educational evaluation (good, regular, poor and very poor) showed that more delta was probably related to a poor evaluation and more alpha in occipital areas to a good one. MONOVA also showed highly significant differences in the absolute power in many leads between children with different evaluations. As children with a poor evaluation very frequently had antecedents of risk factors related to brain damage and were from a low socioeconomic status, and both factors have been shown to affect absolute power, it may be that the differences observed were due to these causes. However, relative power correlated more with the learning problems. Children with minor difficulties, with no antecedents and with good socioeconomic status had more theta in almost all leads than children with a good evaluation and with the same characteristics. Children with a poor, or very poor, evaluation had more delta in left frontal and temporal areas (F3, F7 and T3) which may reflect underlying cerebral dysfunction of these regions directly involved in reading and writing processes.

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