Search

We assessed the diagnostic utility of deamidated gliadin peptide immunoglobulin G (DGP-IgG) in pediatric patients without immunoglobulin A deficiency who underwent tissue transglutaminase immunoglobulin A (TTG-IgA) screening and biopsy.Patients who had TTG-IgA performed in our laboratory had sample frozen over 1.5 y. If a patient underwent biopsy within 6 months of serology, DGP-IgG was performed on frozen sample. All testing was performed on the BioPlex 2200. Biopsies were assigned a modified Marsh-Oberhuber score. The sensitivity, specificity, and positive and negative predictive values were calculated for TTG-IgA and DGP-IgG for values ≥ 15 u/ml, 15-149 u/ml, and ≥ 150 u/ml using biopsy as gold standard.A total of 458 patients were included. Sensitivity and specificity for DGP-IgG ≥ 15 u/ml and Marsh ≥ 2 was 76% and 87.5% and TTG-IgA ≥ 15 was notably higher at 93.3% and 92.2%. Sensitivity and specificity of DGP-IgG were 66% and 88.9% at moderate and 29.3% and 98.4% at high increases. The positive predictive value of DGP-IgG for celiac disease in TTG-IgA negative patients was 2.8%.Our study suggests DGP-IgG does not add significant value in patients screened for celiac disease.

The Interdisciplinary Assessment Model was created out of the need for improved assessment of learning, especially the assessment of critical thinking and problem solving. Development of the model required establishing a common language for library media specialists to cover various curriculum areas, a comparison of current state standards for subject areas, and creation of rubrics for each stage of the assessment model. The model is based on the "Big Six" model for information problem-solving by Eisenberg and Berkowitz and is derived from an analysis of Kansas content standards for language arts, social studies, mathematics, science, reading, and library media. The model divides student assignments in these six subject areas into five parts, using terminology from the standards for each subject. Rubrics were developed for each of the five parts of an assignment. This paper describes the development of the model, elements of the model, preliminary findings of the current research project which tests the model, and implications for implementing the model in schools. The model facilitated student learning in all grade levels studied and for units of any length. The integrated assessment model is an effective and efficient planning and teaching tool for library media specialists and teachers. Effective implementation requires instruction and tools which will enable teachers to change their instructional strategies to more effectively engage the student learner and to provide feedback to the learner. (Author/SWC) ******************************************************************************** Reproductions supplied by EDRS are the best that can be made from the original document. ********************************************************************************

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection elicits an antibody response that targets several viral proteins including spike (S) and nucleocapsid (N); S is the major target of neutralizing antibodies. Here, we assess levels of anti-N binding antibodies and anti-S neutralizing antibodies in unvaccinated children compared with unvaccinated older adults following infection. Specifically, we examine neutralization and anti-N binding by sera collected up to 52 weeks following SARS-CoV-2 infection in children and compare these to a cohort of adults, including older adults, most of whom had mild infections that did not require hospitalization. Neutralizing antibody titers were lower in children than adults early after infection, but by 6 months titers were similar between age groups. The neutralizing activity of the children’s sera decreased modestly from one to six months; a pattern that was not significantly different from that observed in adults. However, infection of children induced much lower levels of anti-N antibodies than in adults, and levels of these anti-N antibodies decreased more rapidly in children than in adults, including older adults. These results highlight age-related differences in the antibody responses to SARS-CoV-2 proteins and, as vaccines for children are introduced, may provide comparator data for the longevity of infection-elicited and vaccination-induced neutralizing antibody responses.

Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection elicits antibodies (Abs) that bind several viral proteins such as the spike entry protein and the abundant nucleocapsid (N) protein. We examined convalescent sera collected through 6 months (~24wks) post-SARS-CoV-2 infection in children to evaluate changes in neutralization potency and N-binding. Methods Outpatient, hospitalized, and community recruited volunteers < 18 years with COVID-19 were enrolled in a longitudinal study at Seattle Children’s Hospital. Analysis includes symptomatic and asymptomatic children with laboratory-confirmed SARS-CoV-2 infection who provided blood samples at approximately 4wks (range: 2-18wks, IQR:4-8wks) and 24 wks (range: 23-35wks, IQR:25-27wks) after diagnosis. We measured neutralizing Ab using an in-house pseudoneutralization assay and anti-N binding Ab using the Abbott Architect assay. Results Of 32 children enrolled between April 2020 and January 2021, 27 had no underlying immunocompromised state and 25 of these 27 children had symptomatic disease. Ten of 27 had a > 2-fold decrease neutralization titers between 4 and 24wks (most were < 10-fold); 12 had < 2-fold change; and 5 had neutralization titers that increased > 2-fold over time (Fig. 1A). All but one of these 27 children had detectable neutralizing activity at 24wks. Anti-N Abs were assessed for 25 children at 4wks and 17 children at 24wks (data pending for 14 samples); all children with paired samples had a > 1.75-fold Abbott index reduction at 24wks, and 5 children had no detectable anti-N Abs by 24wks (Fig. 2A). An additional 5 children with symptomatic disease had complicating immunosuppression or multiple blood transfusions; 2 had decreasing neutralizing titers, 2 increased, and 1 had no change (Fig. 1B). Anti-N Abs were undetectable for one child by 24wks (data pending for 4 samples) (Fig. 2B). No participants received COVID-19 vaccine. Figure 1. Pseusoneutralization titers in children over time. Figure 2. Nucleocapsid-binding antibody titers in children over time. Conclusion We show neutralizing Abs wane to a small degree over 24wks post-SARS-CoV-2 infection and remain detectable in most children. In contrast, anti-N Abs decreased, becoming undetectable in some children by 24wks. These findings add to understanding of the natural history of SARS-CoV-2 immunity in children. * This study was supported by CDC BAA75D301-20-R-67897 Disclosures Jesse Bloom, PhD, Flagship Labs 77 (Consultant)Moderna (Consultant) Janet A. Englund, MD, AstraZeneca (Consultant, Grant/Research Support)GlaxoSmithKline (Research Grant or Support)Meissa Vaccines (Consultant)Pfizer (Research Grant or Support)Sanofi Pasteur (Consultant)Teva Pharmaceuticals (Consultant)

Appropriate laboratory test utilization is of growing interest in the face of rising healthcare costs and documented evidence of over- and under-utilization. Building from published literature, laboratory organizations have recently published guidelines for establishing laboratory utilization management programs. However, systematic reviews and meta-analyses have consistently struggled to define rigorous evidence-based best practice recommendations due to the paucity of published data or the heterogeneity of available data. We sought to gain information about utilization practices and programs currently in use and which factors contribute to their success by distributing a survey among laboratory professionals. The survey received seventy-four eligible respondents. We observed a wide range in the duration of laboratory utilization programs and the number of stewardship initiatives. In addition, there was great variety in the utilization practices used and the tests or processes targeted by programs. There was similarity in how initiatives are evaluated and who is involved with utilization programs. Finally, respondents often credited a multidisciplinary committee, support from leadership, and strong IT support/data access as important factors for their program's perceived success. Many of these factors agree with previously published literature.

Macrophages are innate immune cells that play critical roles in tissue homeostasis, inflammation, and immune oncology. Macrophages differentiated from human induced pluripotent stem cells (iPSCs) overcome many limitations of using peripheral blood derived macrophages. The ability to scale up and cryopreserve a large amount of end stage macrophages from single clonal iPSCs from normal and disease specific donors offers a unique opportunity for genomic analysis and drug screening. The present study describes the step wise generation and characterization of macrophages from iPSCs using a defined serum free method amenable to scale up to generate a large batch of pure end stage cryopreservable macrophages expressing CD68, CD33, CD11c, CD11b, CD1a, HLA-DR, CD86, CD64, CD80, CD206, CD169, CD47, HLA-ABC, and CX3CR. The end stage macrophages pre and post cryopreservation retain purity, morphology, responsiveness to stimuli and display robust phagocytic function coming right out of cryopreservation. The same differentiation process was used to generate end stage macrophages from isogenic iPSCs engineered to mimic mutations associated with Parkinson’s disease (SNCA A53T), neuronal ceroid lipofuscinosis (GRN2/GRN R493X), and Rett syndrome (MECP2-Knockout). End stage macrophages from isogenic engineered clones displayed differential macrophage-specific purity markers, phagocytic function, and response to specific stimuli. Thus, generating a panel of functional, physiologically relevant iPSC-derived macrophages can potentially facilitate the understanding of neural inflammatory responses associated with neurodegeneration.

Design and implement a concurrent campaign of influenza immunization and tuberculosis (TB) screening for health care workers (HCWs) that can reduce the number of clinic visits for each HCW.A discrete-event simulation model was developed to support issues of resource allocation decisions in planning and operations phases.The campaign was compressed to100 days in 2010 and further compressed to 75 days in 2012 and 2013. With more than 5000 HCW arrivals in 2011, 2012 and 2013, the 14-day goal of TB results was achieved for each year and reduced to about 4 days in 2012 and 2013.Implementing a concurrent campaign allows less number of visiting clinics and the compressing of campaign length allows earlier immunization. The support of simulation modelling can provide useful evaluations of different configurations.

Human induced pluripotent stem cells (hiPSCs) have been shown to differentiate along the retinal lineage in a manner that mimics normal mammalian development. Under certain culture conditions, hiPSCs form optic vesicle-like structures (OVs), which contain proliferating progenitors capable of yielding all neural retina (NR) cell types over time. Such observations imply conserved roles for regulators of retinogenesis in hiPSC-derived cultures and the developing embryo. However, whether and to what extent this assumption holds true has remained largely uninvestigated. We examined the role of a key NR transcription factor, visual system homeobox 2 (VSX2), using hiPSCs derived from a patient with microphthalmia caused by an R200Q mutation in the VSX2 homeodomain region. No differences were noted between (R200Q)VSX2 and sibling control hiPSCs prior to OV generation. Thereafter, (R200Q)VSX2 hiPSC-OVs displayed a significant growth deficit compared to control hiPSC-OVs, as well as increased production of retinal pigmented epithelium at the expense of NR cell derivatives. Furthermore, (R200Q)VSX2 hiPSC-OVs failed to produce bipolar cells, a distinctive feature previously observed in Vsx2 mutant mice. (R200Q)VSX2 hiPSC-OVs also demonstrated delayed photoreceptor maturation, which could be overcome via exogenous expression of wild-type VSX2 at early stages of retinal differentiation. Finally, RNAseq analysis on isolated hiPSC-OVs implicated key transcription factors and extracellular signaling pathways as potential downstream effectors of VSX2-mediated gene regulation. Our results establish hiPSC-OVs as versatile model systems to study retinal development at stages not previously accessible in humans and support the bona fide nature of hiPSC-OV-derived retinal progeny. Stem Cells 2014;32:1480–1492

Purpose We sought to determine if human induced pluripotent stem cells (iPSCs) derived from blood could produce optic vesicle-like structures (OVs) with the capacity to stratify and express markers of intercellular communication. Methods Activated T-lymphocytes from a routine peripheral blood sample were reprogrammed by retroviral transduction to iPSCs. The T-lymphocyte-derived iPSCs (TiPSCs) were characterized for pluripotency and differentiated to OVs using our previously published protocol. TiPSC-OVs were then manually isolated, pooled, and cultured en masse to more mature stages of retinogenesis. Throughout this stepwise differentiation process, changes in anterior neural, retinal, and synaptic marker expression were monitored by PCR, immunocytochemistry, and/or flow cytometry. Results TiPSCs generated abundant OVs, which contained a near homogeneous population of proliferating neuroretinal progenitor cells (NRPCs). These NRPCs differentiated into multiple neuroretinal cell types, similar to OV cultures from human embryonic stem cells and fibroblast-derived iPSCs. In addition, portions of some TiPSC-OVs maintained their distinctive neuroepithelial appearance and spontaneously formed primitive laminae, reminiscent of the developing retina. Retinal progeny from TiPSC-OV cultures expressed numerous genes and proteins critical for synaptogenesis and gap junction formation, concomitant with the emergence of glia and the upregulation of thrombospondins in culture. Conclusions We demonstrate for the first time that human blood-derived iPSCs can generate retinal cell types, providing a highly convenient donor cell source for iPSC-based retinal studies. We also show that cultured TiPSC-OVs have the capacity to self-assemble into rudimentary neuroretinal structures and express markers indicative of chemical and electrical synapses.