The quality assurance program presented here provides a means to maximize and maintain the performance of individual flow cytometers in a facility. To optimize performance, we recommend performing ...all three steps (optimization, calibration and standardization) in this program when a new flow cytometer is installed or whenever the flow cytometer's optical path is altered (e.g., lasers, filters or detectors are replaced). The complete process requires 3-4 h. On a more frequent basis, only a subset of these procedures need to be performed as a part of daily maintenance routines. The data generated can be tracked to monitor the instrument and determine whether service is needed. In addition, the data can provide a metric for whether repairs and upgrades have improved or harmed performance, and for future instrument-to-instrument comparisons. In sum, the procedures presented here represent an updated framework for optimizing, calibrating and standardizing a flow cytometer for daily use.
After compensation, the measurement errors arising from multiple fluorescences spilling into each detector become evident by the spreading of nominally negative distributions. Depending on the ...instrument configuration and performance, and reagents used, this “spillover spreading” (SS) affects sensitivity in any given parameter. The degree of SS had been predicted theoretically to increase with measurement error, i.e., by the square root of fluorescence intensity, as well as directly related to the spectral overlap matrix coefficients. We devised a metric to quantify SS between any pair of detectors. This metric is intrinsic, as it is independent of fluorescence intensity. The combination of all such values for one instrument can be represented as a spillover spreading matrix (SSM). Single‐stained controls were used to determine the SSM on multiple instruments over time, and under various conditions of signal quality. SSM values reveal fluorescence spectrum interactions that can limit the sensitivity of a reagent in the presence of brightly‐stained cells on a different color. The SSM was found to be highly reproducible; its non‐trivial values show a CV of less than 30% across a 2‐month time frame. In addition, the SSM is comparable between similarly‐configured instruments; instrument‐specific differences in the SSM reveal underperforming detectors. Quantifying and monitoring the SSM can be a useful tool in instrument quality control to ensure consistent sensitivity and performance. In addition, the SSM is a key element for predicting the performance of multicolor immunofluorescence panels, which will aid in the optimization and development of new panels. We propose that the SSM is a critical component of QA/QC in evaluation of flow cytometer performance. Published 2013 Wiley‐ Periodicals, Inc.
Mixed reality has made its first step towards democratization in 2017 with the launch of a first generation of commercial devices. As a new medium, one of the challenges is to develop interactions ...using its endowed spatial awareness and body tracking. More specifically, at the crossroad between artificial intelligence and human-computer interaction, the goal is to go beyond the Window, Icon, Menu, Pointer (WIMP) paradigm humans are mainly using on desktop computer. Hand interactions either as a standalone modality or as a component of a multimodal modality are one of the most popular and supported techniques across mixed reality prototypes and commercial devices. In this context, this paper presents scoping literature review of hand interactions in mixed reality. The goal of this review is to identify the recent findings on hand interactions about their design and the place of artificial intelligence in their development and behavior. This review resulted in the highlight of the main interaction techniques and their technical requirements between 2017 and 2022 as well as the design of the Metaphor-behavior taxonomy to classify those interactions.
Cellular uptake and intracellular release efficiency of biomacromolecules is low because of hurdles in the cell membrane that result in limited access to intra-cellular targets with few functional ...effects. Cell-penetrating peptides (CPPs) act as cargo delivery vehicles to promote therapeutic molecule translocation. Here, we describe the novel CPP-Dot1l that not only penetrates by itself, but also mediates cargo translocation in cultured cells, as confirmed by fluorescence microscopy and fluorescence spectrophotometry. We conducted cytotoxicity assays and safety evaluations, and determined peptide-membrane interactions to understand the possible pathway for cargo translocation. Additional nucleic acid and covalently conjugated green fluorescence protein (GFP) studies mediated by CPP-Dot1l were conducted to show functional delivery potential. Results indicate that CPP-Dot1l is a novel and effective CPP due to its good penetrating properties in different cell lines and its ability to enter cells in a concentration-dependent manner. Its penetration efficiency can be prompted by DMSO pretreatment. In addition, not only can it mediate plasmid delivery, but CPP-Dot1l can also deliver GFP protein into cytosol. In conclusion, the findings of this study showed CPP-Dot1l is an attractive pharmaceutical and biochemical tool for future drug, regenerative medicine, cell therapy, gene therapy, and gene editing-based therapy development.
Background/aim Low-cost commercial bCPAP devices have been deployed in resource-limited settings to treat neonatal respiratory failure. The use of these devices has increased access to pediatric ...respiratory support for infants. However, constrained resources may result in substitution of recommended consumables and/or use in older age groups. We hypothesized that commercially available bCPAP devices, the standard WHO-style device and various improvised adaptations would all generate effective, safe positive pressure at the patient interface. Methods Performance of 2 commercially available bCPAP devices was tested against the standard WHO-style bCPAP device, as well as several improvised modifications of these devices, by measuring positive pressure delivered at the patient interface. Variables tested included different flow rates, patient interfaces and respiratory circuit tubing. Results Both commercial devices utilized according to manufacturer recommendations generated the expected positive pressure at the patient interface. When testing the recommended WHO-style bCPAP device with recommended materials as well as other improvised modifications, we found variable and potentially unpredictable generation of positive pressure at the patient interface. Conclusions Modified or improvised bCPAP devices should be used with extreme caution as the support provided may be more or less than expected depending on respiratory tubing and flow rates employed. Our data support the effectiveness of bCPAP in newborns and young infants. But, to our knowledge, there are no bCPAP patient interfaces for older children effective with low liter flow devices. Therefore, based on these results, we recommend against using WHO-style bCPAP devices for non-infant patients with respiratory failure and instead recommend using standard oxygen therapy with nasal cannulae or face-masks, as well as early consideration of transfer to a higher level of care. Keywords: Pediatrics, Bubble CPAP, Critical care, Global health, Resource limited setting; respiratory failure
The simian immunodeficiency virus (SIV) challenge model of lentiviral infection is often used as a model to human immunodeficiency virus type 1 (HIV-1) for studying vaccine mediated and immune ...correlates of protection. However, knowledge of the structure of the SIV envelope (Env) glycoprotein is limited, as is knowledge of binding specificity, function and potential efficacy of SIV antibody responses. In this study we describe the use of a competitive probe binding sort strategy as well as scaffolded probes for targeted isolation of SIV Env-specific monoclonal antibodies (mAbs). We isolated nearly 70 SIV-specific mAbs directed against major sites of SIV Env vulnerability analogous to broadly neutralizing antibody (bnAb) targets of HIV-1, namely, the CD4 binding site (CD4bs), CD4-induced (CD4i)-site, peptide epitopes in variable loops 1, 2 and 3 (V1, V2, V3) and potentially glycan targets of SIV Env. The range of SIV mAbs isolated includes those exhibiting varying degrees of neutralization breadth and potency as well as others that demonstrated binding but not neutralization. Several SIV mAbs displayed broad and potent neutralization of a diverse panel of 20 SIV viral isolates with some also neutralizing HIV-2(7312A). This extensive panel of SIV mAbs will facilitate more effective use of the SIV non-human primate (NHP) model for understanding the variables in development of a HIV vaccine or immunotherapy.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Both SIV and SHIV are powerful tools for evaluating antibody-mediated prevention and treatment of HIV-1. However, owing to a lack of rhesus-derived SIV broadly neutralizing antibodies (bnAbs), ...testing of bnAbs for HIV-1 prevention or treatment has thus far been performed exclusively in the SHIV NHP model using bnAbs from HIV-1-infected individuals. Here we describe the isolation and characterization of multiple rhesus-derived SIV bnAbs capable of neutralizing most isolates of SIV. Eight antibodies belonging to two clonal families, ITS102 and ITS103, which target unique epitopes in the CD4 binding site (CD4bs) region, were found to be broadly neutralizing and together neutralized all SIV strains tested. A rare feature of these bnAbs and two additional antibody families, ITS92 and ITS101, which mediate strain-specific neutralizing activity against SIV from sooty mangabeys (SIVsm), was their ability to achieve near complete (i.e. 100%) neutralization of moderately and highly neutralization-resistant SIV. Overall, these newly identified SIV bnAbs highlight the potential for evaluating HIV-1 prophylactic and therapeutic interventions using fully simian, rhesus-derived bnAbs in the SIV NHP model, thereby circumventing issues related to rapid antibody clearance of human-derived antibodies, Fc mismatch and limited genetic diversity of SHIV compared to SIV.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Membrane-damaged cells caused by either mechanical trauma or through normal biological processes can produce artifacts in immunophenotyping analysis by flow cytometry. Dead cells can nonspecifically ...bind monoclonal antibody conjugates, potentially leading to erroneous conclusions, particularly when cell frequencies are low. To date, DNA intercalating dyes (Ethidium monoazaide (EMA), Propidium Iodide, 7AAD, etc.) or Annexin V have been commonly used to exclude dead cells; however, each suffer from technical problems. The first issue with such dyes is the dependence on a consistent dead cell source for fluorescence compensation. Another major issue is the stability of the staining; except for EMA, fixation and permeablization used for intracellular staining procedures can cause loss of fluorescence. EMA requires a UV exposure to covalently bond to DNA; while this dye is effective and is not affected by intracellular treatments it is weakly fluorescent. Here we report on the optimization of a new class of viability dyes, the amine reactive viability dyes (ViD) as a dead cell exclusion marker. The inclusion of ViD into the staining panel was found to be simple, reproducible and can have a significant benefit on the accuracy of identifying appropriate cell populations. We show the fluorescence of cells stained with these dyes correlates with traditional dead cell discriminating markers, even after fixation and permeabilization. Amine reactive viability dyes are a powerful tool for fluorescence immunophenotyping experiments.
Obesity is the most common chronic disease in the United States today. Additional therapies are needed to improve obesity treatment.
A swallowable, gas-filled intragastric balloon system was approved ...for the treatment of obesity by Food and Drug Administration in September 2016 and commercialization started January 2017. A registry was made available to physicians to capture evidence of safety and effectiveness with use.
United States private clinics, surgery centers, and hospitals.
This study is a retrospective analysis of a prospective registry of patients with body mass index (BMI) ≥25 kg/m
that initiated therapy in the first year. Data on demographics, procedural timing, weight loss, adverse events, and device deficiencies were captured.
The final analysis comprised 1343 patients across 108 treating physicians (mean age 45.7 ± 10.8 yr, 78.6% female, baseline BMI of 35.4 ± 5.4 kg/m
). Nonserious and serious adverse events were reported in 14.2% and .15% of patients, respectively. There were 7 balloon deflations, none caused obstruction. Weight loss in the indicated use (BMI 30-40 kg/m
) was 9.7 ± 6.1 kg and 10.0 ± 6.1% total body weight loss (TBWL). Weight loss in other BMI categories was 8.2 ± 5.6 kg or 10.3 ± 7.0% total body weight loss for BMI 25 to 29.9 kg/m
and 11.6 ± 7.8 kg or percent total body weight loss 9.3 ± 6.0 for BMI >40 kg/m
.
This swallowable gas-filled intragastric balloon system is safe and effective at inducing weight loss and offers physicians another tool for patients whose obesity has been resistant to noninvasive treatments.
► A new portable sensory based tactile device is described. ► The portable device is capable of stimulating 4 finger tips independently which allows for the study of cortical–cortical interactions ...and their direct impact on sensory percepts. ► Conditioning stimuli have a profound, yet predictable, impact on simple tactile tests in healthy individuals.
Current methods for applying multi-site vibratory stimuli to the skin typically involve the use of multiple, individual vibrotactile stimulators. Limitations of such an arrangement include difficulty with both positioning the stimuli as well as ensuring that stimuli are delivered in a synchronized and deliberate manner. Previously, we reported a two-site tactile stimulator that was developed in order to solve these problems (Tannan et al., 2007a). Due to both the success of that novel stimulator and the limitations that were inherent in that device, we designed and fabricated a four-site stimulator that provides a number of advantages over the previous version. First, the device can stimulate four independent skin sites and is primarily designed for stimulating the digit tips. Second, the positioning of the probe tips has been re-designed to provide better ergonomic hand placement. Third, the device is much more portable than the previously reported stimulator. Fourth, the stimulator head has a much smaller footprint on the table or surface where it resides. To demonstrate the capacity of the device for delivering tactile stimulation at four independent sites, a finger agnosia protocol, in the presence and absence of conditioning stimuli, was conducted on seventeen healthy control subjects. The study demonstrated that with increasing amplitudes of vibrotactile conditioning stimuli concurrent with the agnosia test, inaccuracies of digit identification increased, particularly at digits D3 and D4. The results are consistent with prior studies that implicated synchronization of adjacent and near-adjacent cortical ensembles with conditioning stimuli in impacting TOJ performance (Tommerdahl et al., 2007a,b).
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