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The combination of 18 F-Sodium Fluoride (18 F-NaF) and 18 F-FluoroDeoxyGlucose (18 F-FDG) for positron emission tomography (PET) imaging of the equine foot is appealing for detection of both osseous and soft tissue lesions in a single scan. As the combination of tracers could lead to a loss of information, a sequential approach, consisting in imaging with one tracer prior to injecting the second tracer, might be valuable. The goals of this prospective, methods comparison, exploratory study were to establish the order of tracer injection and timing for imaging. Six research horses were imaged under general anesthesia with 18 F-NaF PET, 18 F-FDG PET, dual 18 F-NaF/18 F-FDG PET, and CT. Proper uptake could be identified in tendon lesions as early as 10 min after 18F-FDG injection. Bone uptake was limited when 18F-NaF was injected under general anesthesia, even at 1 h after injection, when compared with 18 F-NaF injection prior to anesthesia. The sensitivity and specificity of the dual tracer scans were 0.77 (0.63 to 0.86) and 0.98 (0.96 to 0.99) respectively, to assess 18 F-NaF uptake and 0.5 (0.28 to 0.72) and 0.98 (0.95 to 0.99), respectively, for 18F-FDG uptake. These results suggest that the sequential dual tracer approach is a pertinent technique to optimize the PET data gained from a single anesthetic episode. Based on dynamics of tracer uptake, the optimal protocol consists in injecting 18F-NaF prior to anesthesia, acquire 18F-NaF data then inject 18F-FDG and start acquisition of dual tracer PET data 10 min later. This protocol should be further validated in a larger clinical study.
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Fluorodesoxiglucosa F18 , Radiofármacos , Caballos , Animales , Fluoruro de Sodio , Estudios Prospectivos , Tomografía de Emisión de Positrones/veterinaria , Tomografía de Emisión de Positrones/métodosRESUMEN
In tumor cells, ketolysis "via" succinyl-CoA: 3-oxoacid-CoAtransferase (SCOT) and acetyl-CoA acetyltransferase 1 (ACAT1) is a major source of mitochondrial acetyl-CoA. Active ACAT1 tetramers stabilize by tyrosine phosphorylation, which facilitates the SCOT reaction and ketolysis. Tyrosine phosphorylation of pyruvate kinase PK M2 has the opposite effect, stabilizing inactive dimers, while pyruvate dehydrogenase (PDH), which is already inhibited by phosphorylation, is acetylated by ACAT1 and is doubly locked. This closes the glycolytic supply of acetyl-CoA. In addition, since tumor cells must synthesize fatty acids to create new membranes, they automatically turn off the degradation of fatty acids into acetyl-CoA ("via" the malonyl-CoA brake for the fatty acid carnityl transporter). Thus, inhibiting SCOT the specific ketolytic enzyme and ACAT1 should hold back tumor progression. However, tumor cells are still able to take up external acetate and convert it into acetyl-CoA in their cytosol "via" an acetyl-CoA synthetase, which feeds the lipogenic pathway; additionally, inhibiting this enzyme would make it difficult for tumor cells to form new lipid membrane and survive.
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BACKGROUND: The Veterans Health Administration (VA) refers patients to community providers for specialty services not available on-site. However, community-level specialist shortages may impede access to care. OBJECTIVE: Compare gynecologist supply in veterans' county of residence versus at their VA site. DESIGN: We identified women veteran VA patients from fiscal year (FY) 2017 administrative data and assessed availability of a VA gynecologist within 50 miles (hereafter called "local") of veterans' VA homesites (per national VA organizational survey data). For the same cohort, we then assessed community-level gynecologist availability; counties with < 2 gynecologists/10,000 women (per the Area Health Resource File) were "inadequate-supply" counties. We examined the proportion of women veterans with local VA gynecologist availability in counties with inadequate versus adequate gynecologist supply, stratified by individual and VA homesite characteristics. Chi-square tests assessed statistical differences. PARTICIPANTS: All women veteran FY2017 VA primary care users nationally. MAIN MEASURES: Availability of a VA gynecologist within 50 miles of a veteran's VA homesite; county-level "inadequate-supply" of gynecologists. KEY RESULTS: Among 407,482 women, 9% were in gynecologist supply deserts (i.e., lacking local VA gynecologist and living in an inadequate-supply county). The sub-populations with the highest proportions in gynecologist supply deserts were rural residents (24%), those who got their primary care at non-VAMC satellite clinics (13%), those who got their care at a site without a women's clinic (13%), and those with American Indian or Alaska Native (12%), or white (12%) race. Among those in inadequate-supply counties, 59.9% had gynecologists at their local VA; however, 40.1% lacked a local VA gynecologist. CONCLUSIONS: Most veterans living in inadequate-supply counties had local VA gynecology care, reflecting VA's critical role as a safety net provider. However, for those in gynecologist supply deserts, expanded transportation options, modified staffing models, or tele-gynecology hubs may offer solutions to extend VA gynecology capacity.
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Ginecología , Veteranos , Instituciones de Atención Ambulatoria , Femenino , Accesibilidad a los Servicios de Salud , Hospitales de Veteranos , Humanos , Estados Unidos , United States Department of Veterans AffairsRESUMEN
PURPOSE: Exploring synaptic density changes during brain growth is crucial to understanding brain development. Previous studies in nonhuman primates report a rapid increase in synapse number between the late gestational period and the early neonatal period, such that synaptic density approaches adult levels by birth. Prenatal synaptic development may have an enduring impact on postnatal brain development, but precisely how synaptic density changes in utero are unknown because current methods to quantify synaptic density are invasive and require post-mortem brain tissue. METHODS: We used synaptic vesicle glycoprotein 2A (SV2A) positron emission tomography (PET) radioligands [11C]UCB-J and [18F]Syn-VesT-1 to conduct the first assessment of synaptic density in the developing fetal brain in gravid rhesus monkeys. Eight pregnant monkeys were scanned twice during the third trimester at two imaging sites. Fetal post-mortem samples were collected near term in a subset of subjects to quantify SV2A density by Western blot. RESULTS: Image-derived fetal brain SV2A measures increased during the third trimester. SV2A concentrations were greater in subcortical regions than in cortical regions at both gestational ages. Near term, SV2A density was higher in primary motor and visual areas than respective associative regions. Post-mortem quantification of SV2A density was significantly correlated with regional SV2A PET measures. CONCLUSION: While further study is needed to determine the exact relationship of SV2A and synaptic density, the imaging paradigm developed in the current study allows for the effective in vivo study of SV2A development in the fetal brain.
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Encéfalo , Glicoproteínas de Membrana , Proteínas del Tejido Nervioso , Animales , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Macaca mulatta/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Tomografía de Emisión de Positrones/métodosRESUMEN
Objective.This work assessed the relationship between image signal-to-noise ratio (SNR) and total-body noise-equivalent count rate (NECR)-for both non-time-of-flight (TOF) NECR and TOF-NECR-in a long uniform water cylinder and 14 healthy human subjects using the uEXPLORER total-body PET/CT scanner.Approach.A TOF-NEC expression was modified for list-mode PET data, and both the non-TOF NECR and TOF-NECR were compared using datasets from a long uniform water cylinder and 14 human subjects scanned up to 12 h after radiotracer injection.Main results.The TOF-NECR for the uniform water cylinder was found to be linearly proportional to the TOF-reconstructed image SNR2in the range of radioactivity concentrations studied, but not for non-TOF NECR as indicated by the reducedR2value. The results suggest that the use of TOF-NECR to estimate the count rate performance of TOF-enabled PET systems may be more appropriate for predicting the SNR of TOF-reconstructed images.Significance.Image quality in PET is commonly characterized by image SNR and, correspondingly, the NECR. While the use of NECR for predicting image quality in conventional PET systems is well-studied, the relationship between SNR and NECR has not been examined in detail in long axial field-of-view total-body PET systems, especially for human subjects. Furthermore, the current NEMA NU 2-2018 standard does not account for count rate performance gains due to TOF in the NECR evaluation. The relationship between image SNR and total-body NECR in long axial FOV PET was assessed for the first time using the uEXPLORER total-body PET/CT scanner.
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Fluorodesoxiglucosa F18 , Tomografía de Emisión de Positrones , Humanos , Tomografía Computarizada por Tomografía de Emisión de Positrones , Tomografía de Emisión de Positrones/métodos , Relación Señal-Ruido , AguaRESUMEN
ABSTRACT: Autoerotic death is defined as an accidental death that occurs when some type of apparatus is used to enhance sexual stimulation and causes an unintended death. We present the case of an atypical autoerotic death of a 23-year-old man found deceased in a bathroom. An electrical cord with a standard wall plug on one end and exposed wires twisted into loops on the other end was on the floor near his body. Minute black burns were present on each nipple. The outlet was protected by a ground fault circuit interrupter that was demonstrated to be in working order. An electrical consultant concluded that the cause of death could not be electrocution. Ground fault circuit interrupters work by detecting differences in current within a circuit. If a ground fault occurs, the circuit is broken to prevent a fatal electrocution. In the present case, the ground fault circuit interrupter did not shut off because the decedent had wired himself in parallel with the circuit. This, with the other scene findings, indicated this to be a case of atypical autoerotic death. Autoerotic deaths by means other than asphyxiation are rare. This case serves to illustrate the circumstances of an atypical autoerotic death by means of electrocution.
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Trastornos Parafílicos , Adulto , Asfixia/etiología , Humanos , Masculino , Trastornos Parafílicos/complicaciones , Adulto JovenRESUMEN
PURPOSE: Patient attrition from the Veterans Health Administration (VA) health care system could undercut its mission to ensure care for eligible veterans. Attrition of women veterans could exacerbate their minority status and impede systemic efforts to provide high-quality care. We obtained women veterans' perspectives on why they left or continued to use VA health care. METHODS: A sampling frame of new women veteran VA patients was stratified by those who discontinued (attriters) and those who continued (non-attriters) using VA care. Semistructured interviews were conducted from 2017 to 2018. Transcribed interviews were coded for women's decision-making, contexts, and recommendations related to health care use. RESULTS: Fifty-one women veterans (25 attriters and 26 non-attriters) completed interviews. Reasons for attrition included challenging patient care experiences (e.g., provider turnover, claim processing challenges) and the availability of private health insurance. Personal experiences with VA care (e.g., gender-specific care) were impactful in women's decision to use VA. The affordability of VA care was influential for both groups to stay connected to services. More than one-third of women originally categorized as attriters described subsequently reentering or planning to reenter VA care. Suggestions to decrease attrition included increasing outreach, improving access, and continuing to tailor care delivery to women veterans' needs. CONCLUSIONS: Understanding the drivers of patients' decisions to use or not use the VA is critical for the development of strategies to improve retention of current patients and optimize health outcomes for veterans. Women veterans described complex reasons why they left or continued using VA, with cost/affordability playing an important role even in considerations of returning to VA after a long hiatus.
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Veteranos , Atención a la Salud , Femenino , Hospitales de Veteranos , Humanos , Estados Unidos , United States Department of Veterans Affairs , Salud de los Veteranos , Salud de la MujerRESUMEN
Absolute quantification of regional tissue concentration of radioactivity in positron emission tomography (PET) is a critical parameter-of-interest across various clinical and research applications and is affected by a complex interplay of factors including scanner calibration, data corrections, and image reconstruction. The emergence of long axial field-of-view (FOV) PET systems widens the dynamic range accessible to PET and creates new opportunities in reducing scan time and radiation dose, delayed or low radioactivity imaging, as well as kinetic modeling of the entire human. However, these imaging regimes impose challenging conditions for accurate quantification due to constraints from image reconstruction, low count conditions, as well as large and rapidly changing radioactivity distribution across a large axial FOV. We comprehensively evaluated the quantitative accuracy of the uEXPLORER total-body scanner in conditions that encompass existing and potential imaging applications (such as dynamic imaging and ultralow-dose imaging) using a set of total-body specific phantom and human measurements. Through these evaluations we demonstrated a relative count rate accuracy of ±3%-4% using the NEMA NU 2-2018 protocol, an axial uniformity spread of ±3% across the central 90% axial FOV, and a 3% activity bias spread from 17 to 474 MBq18F-FDG in a 210 cm long cylindrical phantom. Region-of-interest quantification spread of 1% was found by simultaneously scanning three NEMA NU 2 image quality phantoms, as well as relatively stable volume-of-interest quantification across 0.2%-100% of total counts through re-sampled datasets. In addition, an activity bias spread of -2% to +1% post-bolus injections in human subjects was found. Larger bias changes during the bolus injection phase in humans indicated the difficulty in providing accurate PET data corrections for complex activity distributions across a large dynamic range. Our results overall indicated that the quantitative performance achieved with the uEXPLORER scanner was uniform across the axial FOV and provided the accuracy necessary to support a wide range of imaging applications.
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Tomografía Computarizada por Tomografía de Emisión de Positrones , Tomografía de Emisión de Positrones , Fluorodesoxiglucosa F18 , Humanos , Fantasmas de Imagen , Tomografía de Emisión de Positrones/métodos , Tomógrafos Computarizados por Rayos XRESUMEN
Increasing meat demand in terms of both quality and quantity in conjunction with feeding a growing population has resulted in regulatory agencies imposing stringent guidelines on meat quality and safety. Objective and accurate rapid non-destructive detection methods and evaluation techniques based on artificial intelligence have become the research hotspot in recent years and have been widely applied in the meat industry. Therefore, this review surveyed the key technologies of non-destructive detection for meat quality, mainly including ultrasonic technology, machine (computer) vision technology, near-infrared spectroscopy technology, hyperspectral technology, Raman spectra technology, and electronic nose/tongue. The technical characteristics and evaluation methods were compared and analyzed; the practical applications of non-destructive detection technologies in meat quality assessment were explored; and the current challenges and future research directions were discussed. The literature presented in this review clearly demonstrate that previous research on non-destructive technologies are of great significance to ensure consumers' urgent demand for high-quality meat by promoting automatic, real-time inspection and quality control in meat production. In the near future, with ever-growing application requirements and research developments, it is a trend to integrate such systems to provide effective solutions for various grain quality evaluation applications.
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X-ray and gamma-ray photons are widely used for imaging but require a mathematical reconstruction step, known as tomography, to produce cross-sectional images from the measured data. Theoretically, the back-to-back annihilation photons produced by positron-electron annihilation can be directly localized in three-dimensional space using time-of-flight information without tomographic reconstruction. However, this has not yet been demonstrated due to the insufficient timing performance of available radiation detectors. Here, we develop techniques based on detecting prompt Cerenkov photons, which when combined with a convolutional neural network for timing estimation resulted in an average timing precision of 32 picoseconds, corresponding to a spatial precision of 4.8 mm. We show this is sufficient to produce cross-sectional images of a positron-emitting radionuclide directly from the detected coincident annihilation photons, without using any tomographic reconstruction algorithm. The reconstruction-free imaging demonstrated here directly localizes positron emission, and frees the design of an imaging system from the geometric and sampling constraints that normally present for tomographic reconstruction.
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BACKGROUND: When an experienced provider opts to leave a healthcare workforce (attrition), there are significant costs, both direct and indirect. Turnover of healthcare providers is underreported and understudied, despite evidence that it negatively impacts care delivery and negatively impacts working conditions for remaining providers. In the Veterans Affairs (VA) healthcare system, attrition of women's health primary care providers (WH-PCPs) threatens a specially trained workforce; it is unknown what factors contribute to, or protect against, their attrition. OBJECTIVE: Based on evidence that clinic environment, adequate support resources, and workload affect provider burnout and intent to leave, we explored if such clinic characteristics predict attrition of WH-PCPs in the VA, to identify protective factors. DESIGN: This analysis drew on two waves of existing national VA survey data to examine predictors of WH-PCP attrition, via logistic regression. PARTICIPANTS: All 2,259 providers from 140 facilities VA-wide who were WH-PCPs on September 30, 2016. MAIN MEASURES: The dependent variable was WH-PCP attrition in the following year. Candidate predictors were clinic environment (working in: a comprehensive women's health center, a limited women's health clinic, a general primary care clinic, or multiple clinic environments), availability of co-located specialty support resources (mental health, social work, clinical pharmacy), provider characteristics (gender, professional degree), and clinic workload (clinic sessions per week). KEY RESULTS: Working exclusively in a comprehensive women's health center uniquely predicted significantly lower risk of WH-PCP attrition (adjusted odds ratio 0.40; CI 0.19-0.86). CONCLUSIONS: A comprehensive women's health center clinical context may promote retention of this specially trained primary care workforce. Exploring potential mechanisms-e.g., shared mission, appropriate support to meet patients' needs, or a cohesive team environment-may inform broader efforts to retain front-line providers.
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Veteranos , Salud de la Mujer , Instituciones de Atención Ambulatoria , Femenino , Personal de Salud , Humanos , Atención Primaria de Salud , Estados Unidos/epidemiología , United States Department of Veterans Affairs , Recursos HumanosRESUMEN
The world's first total-body PET scanner with an axial field of view (AFOV) of 194 cm is now in clinical and research use at our institution. The uEXPLORER PET/CT system is the first commercially available total-body PET scanner. Here we present a detailed physical characterization of this scanner based on National Electrical Manufacturers Association (NEMA) NU 2-2018 along with a new set of measurements devised to appropriately characterize the total-body AFOV. Methods: Sensitivity, count-rate performance, time-of-flight resolution, spatial resolution, and image quality were evaluated following the NEMA NU 2-2018 protocol. Additional measurements of sensitivity and count-rate capabilities more representative of total-body imaging were performed using extended-geometry phantoms based on the world-average human height (â¼165 cm). Lastly, image quality throughout the long AFOV was assessed with the NEMA image quality (IQ) phantom imaged at 5 axial positions and over a range of expected total-body PET imaging conditions (low dose, delayed imaging, short scan duration). Results: Our performance evaluation demonstrated that the scanner provides a very high sensitivity of 174 kcps/MBq, a count-rate performance with a peak noise-equivalent count rate of approximately 2 Mcps for total-body imaging, and good spatial resolution capabilities for human imaging (≤3.0 mm in full width at half maximum near the center of the AFOV). Excellent IQ, excellent contrast recovery, and low noise properties were illustrated across the AFOV in both NEMA IQ phantom evaluations and human imaging examples. Conclusion: In addition to standard NEMA NU 2-2018 characterization, a new set of measurements based on extending NEMA NU 2-2018 phantoms and experiments was devised to characterize the physical performance of the first total-body PET system. The rationale for these extended measurements was evident from differences in sensitivity, count-rate-activity relationships, and noise-equivalent count-rate limits imposed by differences in dead time and randoms fraction between the NEMA NU 2 70-cm phantoms and the more representative total-body imaging phantoms. Overall, the uEXPLORER PET system provides ultra-high sensitivity that supports excellent spatial resolution and IQ throughout the field of view in both phantom and human imaging.
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Tomografía Computarizada por Tomografía de Emisión de Positrones/instrumentación , Imagen de Cuerpo Entero/instrumentación , Humanos , Límite de Detección , Fantasmas de Imagen , Control de Calidad , Factores de TiempoRESUMEN
Iodination, a chemical or enzymatic incorporation of 125I to specific amino acid side chains, is a commonly used method for labeling antibodies with radioisotopes. Commercially available products make iodination of antibodies a simple and quick process. One example, used here and available at Pierce, is the "Iodination bead," or N-chloro-benzenesulfonamide immobilized on nonporous, polystyrene beads.
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Anticuerpos/química , Radioisótopos de Yodo/química , Marcaje Isotópico/métodos , Microesferas , Poliestirenos/química , Sulfonamidas/química , Halogenación , Oxidación-Reducción , BencenosulfonamidasRESUMEN
Many antibody labeling procedures call for a desalting or purification step requiring size-exclusion chromatography (SEC). The method outlined here contains information needed to desalt an antibody conjugate. Similar procedures would be used for ion-exchange chromatography using a gradient of increasing ionic strength. Resins can be purchased in bulk (as in this protocol), or commercially available columns are available.
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Anticuerpos/química , Anticuerpos/aislamiento & purificación , Cromatografía en Gel/métodos , Coloración y Etiquetado/métodos , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/aislamiento & purificación , Cromatografía por Intercambio Iónico/métodos , Dextranos/química , Resinas Sintéticas/químicaRESUMEN
This introduction outlines general strategies for labeling proteins, with an emphasis on methods that are used primarily for labeling antibodies. It covers the specific site of modification, cross-linker options, types of labels, and postlabeling cleanup methodology, along with the advantages and disadvantages of each method. In general, polyclonal antibodies are more versatile and resistant to activity loss than are monoclonal antibodies. Greater care must be taken when labeling monoclonal antibodies to ensure a quality conjugate. The methods outlined here can be adapted for a variety of labels including multiple labels on the same immunoglobulin. The most important consideration when undertaking an antibody labeling experiment is to maintain the activity of the antibody. This is an empirical process and will often require additional experiments to optimize the label of a particular antibody. When successful, these reagents are very useful and adaptable biomolecules. This introduction provides the reader with methods and options for producing a variety of labeled immunological tools.
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Anticuerpos/química , Reactivos de Enlaces Cruzados/química , Inmunoconjugados/química , Coloración y Etiquetado/métodos , Enfermedad de Alzheimer/diagnóstico , Enfermedad de Alzheimer/inmunología , Péptidos beta-Amiloides/química , Péptidos beta-Amiloides/inmunología , Biotina/química , Carbohidratos/química , Cisteína/química , Oro Coloide/química , Humanos , Inmunoconjugados/inmunología , Marcaje Isotópico/métodos , Proteínas Luminiscentes/química , Compuestos de Sulfhidrilo/químicaRESUMEN
To determine the effects of maternal supplementation on the mRNA abundance of genes associated with metabolic function in fetal muscle and liver, pregnant sows (Landrace × Yorkshire; initial body weight (BW) 221.58 ± 33.26 kg; n = 21) fed a complete gestation diet (corn-soybean meal based diet, CSM) were randomly assigned to 1 of 4 isocaloric supplementation treatments: control (CON, 378 g/d CSM, n = 5), sucrose (SUGAR, 255 g/d crystalized sugar, n = 5), cooked ground beef (BEEF, 330 g/d n = 6), or BEEF + SUGAR (B+S, 165 g/d cooked ground beef and 129 g/d crystalized sugar, n = 5), from days 40 to 110 of gestation. Sows were euthanized on day 111 of gestation. Two male and 2 female fetuses of median BW were selected from each litter, and samples of the longissimus dorsi muscle and liver were collected. Relative transcript level was quantified via qPCR with HPRT1 as the reference gene for both muscle and liver samples. The following genes were selected and analyzed in the muscle: IGF1R, IGF2, IGF2R, GYS-1, IRS-1, INSR, SREBP-1C, and LEPR; while the following were analyzed in the liver: IGF2, IGF2R, FBFase, G6PC, PC, PCK1, FGF21, and LIPC. No effect of fetal sex by maternal treatment interaction was observed in mRNA abundance of any of the genes evaluated (P > 0.11). In muscle, the maternal nutritional treatment influenced (P = 0.02) IGF2 mRNA abundance, with B+S and SUGAR fetuses having lower abundance than CON, which was not different from BEEF. Additionally, SREBP-1 mRNA abundance was greater (P < 0.01) for B+S compared with CON, BEEF, or SUGAR fetuses; and females tended (P = 0.06) to have an increased abundance of SREBP-1 than males. In fetal liver, IGF2R mRNA abundance was greater (P = 0.01) for CON and BEEF than SUGAR and B+S; while FBPase mRNA abundance was greater (P = 0.03) for B+S compared with the other groups. In addition, maternal nutritional tended (P = 0.06) to influence LIPC mRNA abundance, with increased abundance in CON compared with SUGAR and B+S. These data indicate limited changes in transcript abundance due to substitution of supplemental sugar by ground beef during mid to late gestation. However, the differential expression of FBPase and SREBP-1c in response to the simultaneous supplementation of sucrose and ground beef warrants further investigations, since these genes may play important roles in determining the offspring susceptibility to metabolic diseases.
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Suplementos Dietéticos/análisis , Desarrollo Fetal/efectos de los fármacos , Factor II del Crecimiento Similar a la Insulina/genética , Carne Roja/análisis , Sacarosa/administración & dosificación , Porcinos/fisiología , Alimentación Animal/análisis , Animales , Peso Corporal/efectos de los fármacos , Dieta/veterinaria , Femenino , Desarrollo Fetal/genética , Hígado/efectos de los fármacos , Masculino , Músculos/efectos de los fármacos , Embarazo , ARN Mensajero/genética , Porcinos/genética , Porcinos/crecimiento & desarrolloRESUMEN
There are many uses for antibodies labeled with metal ions. Most of these methods involve first attaching a metal chelator to the antibody molecule. This is achieved using standard cross-linking chemistry and then adding the desired metal at appropriate concentration and pH. The method described here outlines a basic procedure for creating a lanthanide conjugate. Lanthanide conjugates are used for proximity assays, as MRI contrast agents, or for mass cytometry experiments. Different metals and chelators can be substituted, but the basic procedures are similar.
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Anticuerpos/química , Quelantes/química , Europio/química , Coloración y Etiquetado/métodos , Cloroformo/química , Reactivos de Enlaces Cruzados/química , Concentración de Iones de Hidrógeno , Metales/químicaRESUMEN
Colloidal gold-antibody conjugates are easy to prepare and are an excellent choice for microscopic applications. Colloidal gold is an aqueous suspension of nanometer-sized particles of gold. Typically, chloroauric acid, HAuCl4, is reduced with dilute solutions of sodium citrate, as described here. This will cause the gold to form small aggregates that will associate with proteins. Gold particles of specific sizes can be isolated and differentiated microscopically, allowing these particles to be used for multiple-label experiments. Colloidal gold-labeled antibodies are widely used in electron microscopy (EM), and can be used for light microscopy but require additional steps (silver enhancement).
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Anticuerpos/química , Oro Coloide/química , Coloración y Etiquetado/métodos , Animales , Anticuerpos/ultraestructura , Bovinos , Concentración de Iones de Hidrógeno , Microscopía Electrónica , Albúmina Sérica Bovina/químicaRESUMEN
An experiment was conducted to test the hypothesis that meat products have digestible indispensable amino acid scores (DIAAS) >100 and that various processing methods will increase standardised ileal digestibility (SID) of amino acids (AA) and DIAAS. Nine ileal-cannulated gilts were randomly allotted to a 9 × 8 Youden square design with nine diets and eight 7-d periods. Values for SID of AA and DIAAS for two reference patterns were calculated for salami, bologna, beef jerky, raw ground beef, cooked ground beef and ribeye roast heated to 56, 64 or 72°C. The SID of most AA was not different among salami, bologna, beef jerky and cooked ground beef, but was less (P < 0·05) than the values for raw ground beef. The SID of AA for 56°C ribeye roast was not different from the values for raw ground beef and 72°C ribeye roast, but greater (P < 0·05) than those for 64°C ribeye roast. For older children, adolescents and adults, the DIAAS for all proteins, except cooked ground beef, were >100 and bologna and 64°C ribeye roast had the greatest (P < 0·05) DIAAS. The limiting AA for this age group were sulphur AA (beef jerky), leucine (bologna, raw ground beef and cooked ground beef) and valine (salami and the three ribeye roasts). In conclusion, meat products generally provide high-quality protein with DIAAS >100 regardless of processing. However, overcooking meat may reduce AA digestibility and DIAAS.
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Labeling antibodies with biotin (biotinylation) is a useful and simple technique. Biotin's small size (244 Da) usually has little effect on the biological activity of the protein target. The most common way to biotinylate an antibody is to cross-link a biotin succinimidyl ester to a primary amine. There are many commercially available types of biotin analogs that can be used for labeling. They vary in reactive group chemistry as well as spacer length. For example, a common analog used for biotinylation is the succinimidyl ester of biotin with an aminohexanoic acid spacer (Long Chain or LC-Biotin), utilized here. A PEG spacer of varying length can also be used.