RESUMEN
Polarization along an apico-basolateral axis is a hallmark of epithelial cells and is essential for their selective barrier and transporter functions, as well as for their ability to provide mechanical resiliency to organs. Loss of polarity along this axis perturbs development and is associated with a wide number of diseases. We describe three steps involved in polarization: symmetry breaking, polarity establishment, and polarity maintenance. While the proteins involved in these processes are highly conserved among epithelial tissues and species, the execution of these steps varies widely and is context dependent. We review both theoretical principles underlying these steps and recent work demonstrating how apico-basolateral polarity is established in vivo in different tissues, highlighting how developmental and physiological contexts play major roles in the execution of the epithelial polarity program.
Asunto(s)
Membrana Basal/metabolismo , Polaridad Celular , Células Epiteliales/citología , Epitelio/metabolismo , Animales , Membrana Basal/citología , Comunicación Celular , Matriz Extracelular/metabolismo , Humanos , Proteínas de la Membrana/metabolismo , Transducción de SeñalRESUMEN
Apico-basolateral polarization is essential for epithelial cells to function as selective barriers and transporters, and to provide mechanical resilience to organs. Epithelial polarity is established locally, within individual cells to establish distinct apical, junctional and basolateral domains, and globally, within a tissue where cells coordinately orient their apico-basolateral axes. Using live imaging of endogenously tagged proteins and tissue-specific protein depletion in the Caenorhabditiselegans embryonic intestine, we found that local and global polarity establishment are temporally and genetically separable. Local polarity is initiated prior to global polarity and is robust to perturbation. PAR-3 is required for global polarization across the intestine but local polarity can arise in its absence, as small groups of cells eventually established polarized domains in PAR-3-depleted intestines in a HMR-1 (E-cadherin)-dependent manner. Despite the role of PAR-3 in localizing PKC-3 to the apical surface, we additionally found that PAR-3 and PKC-3/aPKC have distinct roles in the establishment and maintenance of local and global polarity. Taken together, our results indicate that different mechanisms are required for local and global polarity establishment in vivo.
Asunto(s)
Polaridad Celular , Células Epiteliales , Células Epiteliales/metabolismo , Uniones Intercelulares , Mucosa Intestinal , Intestinos , EpitelioRESUMEN
BACKGROUND: Historically, pharmaceutical companies have struggled with trust and brand reputation among key stakeholders and have adopted innovative marketing strategies to reach patients directly and rebuild those relationships. Social media influencers are a popular strategy to influence younger demographics, including Generation Z and millennials. It is common for social media influencers to work in paid partnerships with brands; this is a multibillion-dollar industry. Long have patients been active in online health communities and social media platforms such as Twitter and Instagram, but in recent years, pharmaceutical marketers have noticed the power of patient persuasion and begun to leverage "patient influencers" in brand campaigns. OBJECTIVE: This study aimed to explore how patient influencers communicate health literacy on pharmaceutical medications on social media to their communities of followers. METHODS: A total of 26 in-depth interviews were conducted with patient influencers using a snowball sampling technique. This study is part of a larger project using an interview guide that included a range of topics such as social media practices, logistics of being an influencer, considerations for brand partnerships, and views on the ethical nature of patient influencers. The constructs of the Health Belief Model were used in this study's data analysis: perceived susceptibility, perceived severity, perceived benefits, perceived barriers, cues to action, and self-efficacy. This study was approved by the institutional review board of the University of Colorado and adhered to ethical standards in interview practice. RESULTS: As patient influencers are a new phenomenon, it was our goal to identify how health literacy on prescription medications and pharmaceuticals is being communicated on social media. Using the constructs of the Health Belief Model to guide the analysis, 3 themes were identified: understanding disease through experience, staying informed on the science or field, and suggesting that physicians know best. CONCLUSIONS: Patients are actively exchanging health information on social media channels and connecting with other patients who share similar diagnoses. Patient influencers share their knowledge and experience in efforts to help other patients learn about disease self-management and improve their quality of life. Similar to traditional direct-to-consumer advertising, the phenomenon of patient influencers raises ethical questions that need more investigation. In a way, patient influencers are health education agents who may also share prescription medication or pharmaceutical information. They can break down complex health information based on expertise and experience and mitigate the loneliness and isolation that other patients may feel without the support of a community.
Asunto(s)
Alfabetización en Salud , Medicamentos bajo Prescripción , Medios de Comunicación Sociales , Humanos , Calidad de Vida , PrescripcionesRESUMEN
Acetylcholinesterase (AChE) is crucial for degrading acetylcholine at cholinergic synapses. In vitro studies suggest that, in addition to its role in nervous system signaling, AChE can also modulate non-neuronal cell properties, although it remains controversial whether AChE functions in this capacity in vivo Here, we show that AChE plays an essential non-classical role in vertebrate gut morphogenesis. Exposure of Xenopus embryos to AChE-inhibiting chemicals results in severe defects in intestinal development. Tissue-targeted loss-of-function assays (via microinjection of antisense morpholino or CRISPR-Cas9) confirm that AChE is specifically required in the gut endoderm tissue, a non-neuronal cell population, where it mediates adhesion to fibronectin and regulates cell rearrangement events that drive gut lengthening and digestive epithelial morphogenesis. Notably, the classical esterase activity of AChE is dispensable for this activity. As AChE is deeply conserved, widely expressed outside of the nervous system, and the target of many environmental chemicals, these results have wide-reaching implications for development and toxicology.
Asunto(s)
Acetilcolinesterasa/metabolismo , Organogénesis/fisiología , Acetilcolina/metabolismo , Acetilcolinesterasa/genética , Animales , Adhesión Celular/fisiología , Embrión no Mamífero/metabolismo , Endodermo/citología , Endodermo/metabolismo , Fibronectinas/genética , Fibronectinas/metabolismo , Inmunohistoquímica , Sistema Nervioso/embriología , Sistema Nervioso/metabolismo , Organogénesis/genética , ARN Mensajero/genética , Xenopus laevis/embriología , Xenopus laevis/genética , Xenopus laevis/metabolismoRESUMEN
The digestive system comprises numerous cells, tissues and organs that are essential for the proper assimilation of nutrients and energy. Many aspects of digestive organ function are highly conserved among vertebrates, yet the final anatomical configuration of the gut varies widely between species, especially those with different diets. Improved understanding of the complex molecular and cellular events that orchestrate digestive organ development is pertinent to many areas of biology and medicine, including the regeneration or replacement of diseased organs, the etiology of digestive organ birth defects, and the evolution of specialized features of digestive anatomy. In this review, we highlight specific examples of how investigations using Xenopus laevis frog embryos have revealed insight into the molecular and cellular dynamics of digestive organ patterning and morphogenesis that would have been difficult to obtain in other animal models. Additionally, we discuss recent studies of gut development in non-model frog species with unique feeding strategies, such as Lepidobatrachus laevis and Eleutherodactylous coqui, which are beginning to provide glimpses of the evolutionary mechanisms that may generate morphological variation in the digestive tract. The unparalleled experimental versatility of frog embryos make them excellent, integrative models for studying digestive organ development across multiple disciplines.
Asunto(s)
Sistema Digestivo/embriología , Xenopus laevis/embriología , Animales , Evolución Biológica , Señalización del Calcio , Comunicación Celular , Sistema Digestivo/citología , Endodermo/citología , Endodermo/embriología , Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Humanos , Morfogénesis , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismoRESUMEN
Few studies have assessed the impact of regulatory actions on air quality improvement through a comprehensive monitoring effort. In this study, we designed saturation sampling of nitrogen oxides (NOX) for the counties of Los Angeles and Alameda (San Francisco Bay) before (2003-2007) and after (2008-2013) implementation of goods movement actions in California. We further separated the research regions into three location categories, including goods movement corridors (GMCs), nongoods movement corridors (NGMCs), and control areas (CTRLs). Linear mixed models were developed to identify whether reductions in NOX were greater in GMCs than in other areas, after controlling for potential confounding, including weather conditions (e.g., wind speed and temperature) and season of sampling. We also considered factors that might confound the relationship, including traffic and cargo volumes that may have changed due to economic downturn impacts. Compared to the pre-policy period, we found reductions of average pollutant concentrations for nitrogen dioxide (NO2) and NOX in GMCs of 6.4 and 21.7 ppb. The reductions were smaller in NGMCs (5.9 and 16.3 ppb, respectively) and in CTRLs (4.6 and 12.1 ppb, respectively). After controlling for potential confounding from weather conditions, season of sampling, and the economic downturn in 2008, the linear mixed models demonstrated that reductions in NO2 and NOX were significantly greater in GMCs compared to reductions observed in CTRLs; there were no statistically significant differences between NGMCs and CTRLs. These results indicate that policies regulating goods movement are achieving the desired outcome of improving air quality for the state, particularly in goods movement corridors where most disadvantaged communities live.
Asunto(s)
Contaminantes Atmosféricos/análisis , Contaminación del Aire/análisis , Comercio/legislación & jurisprudencia , Transportes/legislación & jurisprudencia , California , Monitoreo del Ambiente , Modelos Lineales , Dióxido de Nitrógeno/análisis , Óxidos de Nitrógeno/análisis , Material ParticuladoRESUMEN
Epithelia are tissues with diverse morphologies and functions across metazoans, ranging from vast cell sheets encasing internal organs to internal tubes facilitating nutrient uptake, all of which require establishment of apical-basolateral polarity axes. While all epithelia tend to polarize the same components, how these components are deployed to drive polarization is largely context-dependent and likely shaped by tissue-specific differences in development and ultimate functions of polarizing primordia. The nematode Caenorhabditis elegans (C. elegans) offers exceptional imaging and genetic tools and possesses unique epithelia with well-described origins and roles, making it an excellent model to investigate polarity mechanisms. In this review, we highlight the interplay between epithelial polarization, development, and function by describing symmetry breaking and polarity establishment in a particularly well-characterized epithelium, the C. elegans intestine. We compare intestinal polarization to polarity programs in two other C. elegans epithelia, the pharynx and epidermis, correlating divergent mechanisms with tissue-specific differences in geometry, embryonic environment, and function. Together, we emphasize the importance of investigating polarization mechanisms against the backdrop of tissue-specific contexts, while also underscoring the benefits of cross-tissue comparisons of polarity.
Asunto(s)
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Intestinos , Epitelio , Morfogénesis , Polaridad Celular , Células EpitelialesRESUMEN
Tissue-wide patterning is essential to multicellular development, requiring cells to individually generate polarity axes and coordinate them in space and time with neighbors. Using the C. elegans intestinal epithelium, we identified a patterning mechanism that is informed by cell contact lifetime asymmetry and executed via the scaffolding protein PAR-3 and the transmembrane protein E-cadherin/HMR-1. Intestinal cells break symmetry as PAR-3 and HMR-1 recruit apical determinants into punctate "local polarity complexes" (LPCs) at homotypic contacts. LPCs undergo an HMR-1-based migration to a common midline, thereby establishing tissue-wide polarity. Thus, symmetry breaking results from PAR-3-dependent intracellular polarization coupled to HMR-1-based tissue-level communication, which occurs through a non-adhesive signaling role for HMR-1. Differential lifetimes between homotypic and heterotypic cell contacts are created by neighbor exchanges and oriented divisions, patterning where LPCs perdure and thereby breaking symmetry. These cues offer a logical and likely conserved framework for how epithelia without obvious molecular asymmetries can polarize.
Asunto(s)
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animales , Cadherinas/metabolismo , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Polaridad Celular , Epitelio/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de SeñalRESUMEN
BACKGROUND: HIV spread continues at high rates from infected persons to their sexual partners. In 2009, an estimated 2.6 million new infections occurred globally. People living with HIV (PLHIV) receiving treatment are in contact with health workers and therefore exposed to prevention messages. By contrast, PLHIV not receiving ART often fall outside the ambit of prevention programs. There is little information on their sexual risk behaviors. This study in Mombasa Kenya therefore explored sexual behaviors of PLHIV not receiving any HIV treatment. RESULTS: Using modified targeted snowball sampling, 698 PLHIV were recruited through community health workers and HIV-positive peer counsellors. Of the 59.2% sexually-active PLHIV, 24.5% reported multiple sexual partners. Of all sexual partners, 10.2% were HIV negative, while 74.5% were of unknown HIV status. Overall, unprotected sex occurred in 52% of sexual partnerships; notably with 32% of HIV-negative partners and 54% of partners of unknown HIV status in the last 6 months. Multivariate analysis, controlling for intra-client clustering, showed non-disclosure of HIV status (AOR: 2.38, 95%CI: 1.47-3.84, p < 0.001); experiencing moderate levels of perceived stigma (AOR: 2.94, 95%CI: 1.50-5.75, p = 0.002); and believing condoms reduce sexual pleasure (AOR: 2.81, 95%CI: 1.60-4.91, p < 0.001) were independently associated with unsafe sex. Unsafe sex was also higher in those using contraceptive methods other than condoms (AOR: 5.47, 95%CI: 2.57-11.65, p < 0.001); or no method (AOR: 3.99, 95%CI: 2.06-7.75, p < 0.001), compared to condom users. CONCLUSIONS: High-risk sexual behaviors are common among PLHIV not accessing treatment services, raising the risk of HIV transmission to discordant partners. This population can be identified and reached in the community. Prevention programs need to urgently bring this population into the ambit of prevention and care services. Moreover, beginning HIV treatment earlier might assist in bringing this group into contact with providers and HIV prevention services, and in reducing risk behaviors.
RESUMEN
Sustained polarity and adhesion of epithelial cells is essential for the protection of our organs and bodies, and this epithelial integrity emerges during organ development amidst numerous programmed morphogenetic assaults. Using the developing Caenorhabditis elegans intestine as an in vivo model, we investigated how epithelia maintain their integrity through cell division and elongation to build a functional tube. Live imaging revealed that apical PAR complex proteins PAR-6/Par6 and PKC-3/aPkc remained apical during mitosis while apical microtubules and microtubule-organizing center (MTOC) proteins were transiently removed. Intestine-specific depletion of PAR-6, PKC-3, and the aPkc regulator CDC-42/Cdc42 caused persistent gaps in the apical MTOC as well as in other apical and junctional proteins after cell division and in non-dividing cells that elongated. Upon hatching, gaps coincided with luminal constrictions that blocked food, and larvae arrested and died. Thus, the apical PAR complex maintains apical and junctional continuity to construct a functional intestinal tube.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Células Epiteliales , Mucosa Intestinal , Animales , Caenorhabditis elegans , Células Epiteliales/citología , Células Epiteliales/metabolismo , Mucosa Intestinal/citología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/fisiología , Intestinos/citología , Intestinos/crecimiento & desarrollo , Larva/crecimiento & desarrollo , Centro Organizador de los Microtúbulos/metabolismo , Proteína Quinasa C/metabolismoRESUMEN
Microtubules are polarized intracellular polymers that play key roles in the cell, including in transport, polarity, and cell division. Across eukaryotic cell types, microtubules adopt diverse intracellular organization to accommodate these distinct functions coordinated by specific cellular sites called microtubule-organizing centers (MTOCs). Over 50 years of research on MTOC biology has focused mainly on the centrosome; however, most differentiated cells employ non-centrosomal MTOCs (ncMTOCs) to organize their microtubules into diverse arrays, which are critical to cell function. To identify essential ncMTOC components, we developed the biotin ligase-based, proximity-labeling approach TurboID for use in C. elegans. We identified proteins proximal to the microtubule minus end protein PTRN-1/Patronin at the apical ncMTOC of intestinal epithelial cells, focusing on two conserved proteins: spectraplakin protein VAB-10B/MACF1 and WDR-62, a protein we identify as homologous to vertebrate primary microcephaly disease protein WDR62. VAB-10B and WDR-62 do not associate with the centrosome and instead specifically regulate non-centrosomal microtubules and the apical targeting of microtubule minus-end proteins. Depletion of VAB-10B resulted in microtubule mislocalization and delayed localization of a microtubule nucleation complex ɣ-tubulin ring complex (γ-TuRC), while loss of WDR-62 decreased the number of dynamic microtubules and abolished γ-TuRC localization. This regulation occurs downstream of cell polarity and in conjunction with actin. As this is the first report for non-centrosomal roles of WDR62 family proteins, we expand the basic cell biological roles of this important disease protein. Our studies identify essential ncMTOC components and suggest a division of labor where microtubule growth and localization are distinctly regulated.
Asunto(s)
Caenorhabditis elegans , Centro Organizador de los Microtúbulos , Microtúbulos , Animales , Centrosoma , Proteínas del Citoesqueleto , Proteínas Asociadas a Microtúbulos , Tubulina (Proteína)RESUMEN
Recruiting and retaining an adequate sample is critical to the success of any research project involving humans. Recent reports indicate that the Health Insurance Portability and Accountability Act (HIPAA) privacy rule has adversely affected research. Few resources are available to help researchers navigate the challenges to recruitment and retention after HIPAA privacy rule implementation. This article addresses obstacles to recruitment in prospective clinical research studies related to the HIPAA privacy rule, as well as HIPAA-compliant strategies to enhance recruitment and retention. Recruitment challenges discussed include evolving interpretations of the HIPAA regulations, inability to directly contact potential participants, complexity of HIPAA-required documents, increased costs of recruitment, and an expanding administrative burden. Among the strategies addressed are preparatory research reviews, using clinical collaborators and staff liaisons, prescreening potential participants, minimizing participant burden during the consent process, enhancing participant follow-up, facilitating recruitment for future studies, and streamlining compliance training for staff.
Asunto(s)
Investigación Biomédica/organización & administración , Confidencialidad , Health Insurance Portability and Accountability Act/organización & administración , Selección de Paciente , Proyectos de Investigación , Publicidad , Investigación en Enfermería Clínica/organización & administración , Confidencialidad/legislación & jurisprudencia , Confidencialidad/psicología , Formularios de Consentimiento/organización & administración , Experimentación Humana/legislación & jurisprudencia , Humanos , Consentimiento Informado/legislación & jurisprudencia , Consentimiento Informado/psicología , Medios de Comunicación de Masas , Folletos , Estudios Prospectivos , Proyectos de Investigación/legislación & jurisprudencia , Investigadores/educación , Investigadores/organización & administración , Sujetos de Investigación/economía , Sujetos de Investigación/legislación & jurisprudencia , Sujetos de Investigación/psicología , Estados UnidosRESUMEN
Sub-individual biomarkers are sub-lethal biological responses commonly used in the assessment of wildlife exposure to environmental contaminants. In this study, we examined the activity of glutathione-s-transferase (GST) and lactate dehydrogenase (LDH), and metallothionein (MT) concentrations among captive-raised alligator hatchlings, wild-caught juveniles, and wild-caught adults. Juveniles and adults were collected from three locations in Florida (USA) with varying degrees of contamination (i.e. Lake Apopka (organochlorine polluted site), Merritt Island National Wildlife Refuge (NWR) (metal polluted site), and Lake Woodruff NWR (reference site)). We examined whether changes in the response of these three biomarkers were age and sex dependent or reflected site-specific variations of environmental contaminants. Juvenile alligators from Merritt Island NWR had higher MT concentrations and lower GST activity compared to those from the other two sites. This outcome was consistent with higher metal pollution at this location. Sexually dimorphic patterns of MT and GST (F > M) were observed in juvenile alligators from all sites, although this pattern was not observed in adults. GST activity was lower in captive-raised alligators from Lake Apopka and Merritt Island NWR as compared to animals from Lake Woodruff NWR, suggesting a possible developmental modulator at these sites. No clear patterns were observed in LDH activity. We concluded that GST and MT demonstrate age and sex specific patterns in the alligators inhabiting these study sites and that the observed variation among sites could be due to differences in contaminant exposure.
Asunto(s)
Caimanes y Cocodrilos/metabolismo , Monitoreo del Ambiente/métodos , Hígado/efectos de los fármacos , Contaminantes Químicos del Agua/análisis , Factores de Edad , Caimanes y Cocodrilos/crecimiento & desarrollo , Animales , Biomarcadores/metabolismo , Femenino , Florida , Glutatión Transferasa/metabolismo , L-Lactato Deshidrogenasa/metabolismo , Lagos/química , Hígado/enzimología , Hígado/metabolismo , Masculino , Metalotioneína/metabolismo , Caracteres Sexuales , Contaminantes Químicos del Agua/toxicidadRESUMEN
Epidemiological studies investigating relationships between environmental exposures from air pollution and health typically use residential addresses as a single point for exposure, while environmental exposures in transit, at work, school or other locations are largely ignored. Personal exposure monitors measure individuals' exposures over time; however, current personal monitors are intrusive and cannot be operated at a large scale over an extended period of time (e.g., for a continuous three months) and can be very costly. In addition, spatial locations typically cannot be identified when only personal monitors are used. In this paper, we piloted a study that applied momentary location tracking services supplied by smart phones to identify an individual's location in space-time for three consecutive months (April 28 to July 28, 2013) using available Wi-Fi networks. Individual exposures in space-time to the traffic-related pollutants Nitrogen Oxides (NOX) were estimated by superimposing an annual mean NOX concentration surface modeled using the Land Use Regression (LUR) modeling technique. Individual's exposures were assigned to stationary (including home, work and other stationary locations) and in-transit (including commute and other travel) locations. For the individual, whose home/work addresses were known and the commute route was fixed, it was found that 95.3% of the time, the individual could be accurately identified in space-time. The ambient concentration estimated at the home location was 21.01 ppb. When indoor/outdoor infiltration, indoor sources of air pollution and time spent outdoors were taken into consideration, the individual's cumulative exposures were 28.59 ppb and 96.49 ppb, assuming a respective indoor/outdoor ratio of 1.33 and 5.00. Integrating momentary location tracking services with fixed-site field monitoring, plus indoor-outdoor air exchange calibration, makes exposure assessment of a very large population over an extended time period feasible.
Asunto(s)
Contaminantes Atmosféricos/análisis , Contaminación del Aire/estadística & datos numéricos , Teléfono Celular , Exposición a Riesgos Ambientales/estadística & datos numéricos , Monitoreo del Ambiente/instrumentación , Monitoreo del Ambiente/métodos , HumanosRESUMEN
Increasing diabetes prevalence has been found to be a primary driver of increased health care costs in the United States. This policy brief examines the impact of diabetes on hospitalizations and related hospitalization costs in California. Using 2011 hospital patient discharge data and annual financial data from the Office of Statewide Health Planning and Development (OSHPD), this study found that patients with diabetes represented 31 percent of hospitalizations in California in 2011 among patients 35 years or older, including 39 percent of African-American and Asian-American patients and 43 percent of Latino patients. Moreover, these hospitalizations cost nearly $2,200 more per hospitalization than those for patients without diabetes, regardless of the primary reason for the hospitalization. Given that approximately 90-95 percent of diagnosed diabetes among adults is type 2 diabetes and is therefore preventable, public health measures can and should be taken to relieve the burden of type 2 diabetes. Such measures include promoting a healthy diet and regular physical activity and providing adequate access to primary and specialty care.
Asunto(s)
Diabetes Mellitus Tipo 2/epidemiología , Costos de la Atención en Salud/tendencias , Hospitalización/economía , Adulto , California , Costo de Enfermedad , Diabetes Mellitus Tipo 2/prevención & control , Dieta , Etnicidad/estadística & datos numéricos , Ejercicio Físico , Predicción , Promoción de la Salud , Accesibilidad a los Servicios de Salud , Encuestas Epidemiológicas , Hospitalización/estadística & datos numéricos , Humanos , Prevalencia , Atención Primaria de Salud , Estados UnidosRESUMEN
The magnitude and functional phenotype (e.g. proliferation, immune stimulation) of vaccine-induced T-cell responses are likely to be critical in defining responses that can control pathogenic challenge. Current multi-parameter flow cytometric techniques may not be sufficient to measure all of these different functions, since characterizing T-cell responses by flow cytometry is presently limited to concurrent measurement of at most 10 cytokines/chemokines. Here, we describe extensive studies conducted using standardized GCLP procedures to optimize and qualitatively/quantitatively qualify a multiplex bead array (MBA) performed on supernatant collected from stimulated peripheral blood mononuclear cells (PBMC) to assess 12 cytokines and chemokines of interest. Our optimized MBA shows good precision (intra-assay, inter-day, inter-technician; coefficients of variation <30%) and linearity for most of the analytes studied. We also developed positivity criteria that allow us to define a response as positive or negative with a high degree of confidence. In conclusion, we provide a detailed description of the qualification of an MBA, which permits quantitative and qualitative evaluation of vaccine-induced immunogenicity and analysis of immune correlates of protection. This assay provides an excellent complement to the existing repertoire of assays for assessing immunogenicity in HIV vaccine clinical trials.