bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Unique Multiplex Assay to Identify Th1/Th2 Cytokine Profile in MS Patients
Authors: Madiraju Charitha , Karp Jacob Alexander , Oppong Martha , Matson Robert 
Published: January 2023

Abstract
Multiple Sclerosis (MS) is a chronic neuroinflammatory, demyelinating autoimmune disease of the central nervous system (CNS). There are diverse views on the paradigm that disease pathogenesis of MS is due to an imbalance in the proinflammatory T helper 1 (Th1)-type cytokines and anti-inflammatory Th2-type cytokines. Several factors play a crucial role in Th1/Th2 polarization that contributes towards disease activation or progression and this is dependent on MS disease sub-type. The goal of our research study is to investigate a unique and robust multiplex assay for identification of Th1 versus Th2 cytokine expression pattern as applicable to samples derived from MS patients. Towards this end, a sandwich ELISA-based multiplex assay method in a 96-well plate format is designed for establishing Th1/Th2 cytokine profile. Cerebrospinal fluid (CSF) samples derived from MS patients diagnosed with either Relapsing-Remitting MS (RRMS, N = 10) or Secondary progressive MS (SPMS, N = 10) and two unaffected control CSF samples derived from the otherwise healthy human subjects are included in the study. Multiplex ELISA analysis in a 96-well format included TNF, IFN-gamma, IL-4, IL-13 and IL-17E cytokines. Each well is imprinted in a 3*3 format with a capture antibody against a given cytokine of interest. The colorimetric read-outs from multiplex assay are analyzed by ImageJ. Preliminary results identify the multiplex assay to be robust, unique and cost-effective for establishing Th1/Th2 cytokine profiles in two different sub-types of MS. Th1/Th2 cytokine pattern could help predict the course of MS disease progression and remission.
Linking human health effects to water quality using a rapid, noninvasive salivary-IgG antibody multiplex immunoassay
Authors: Augustine S. Jason , Eason Tarsha N , Griffin Shannon M , Simmons Kaneatra J , Sams Elizabeth A , Oshima Kevin H , Dufour Al , Wade Timothy J 
Published: January 2023

Abstract
Understanding and curbing the mounting global increase in waterborne pathogen outbreaks are quickly becoming major priorities for public health professionals and policy makers. According to the World Health Organization (WHO), 60% of global diarrheal deaths are caused by unsafe water and lack of sanitation or hygiene. The WHO estimates that at least 2 billion people use a drinking water source contaminated with feces. Contaminated water -- shown to transmit diseases such as cholera, dysentery, typhoid, hepatitis A, and polio -- is estimated to cause almost half a million diarrheal deaths each year. The United States Environmental Protection Agency has prioritized efforts to understand the links between water quality and human health effects. To that end, we have developed and applied a salivary-IgG antibody multiplex immunoassay to measure human exposures and associated health effects to multiple pathogens simultaneously. Saliva is emerging as a cost-effective, noninvasive biofluid that is well-accepted by children. The multiplex immunoassay has afforded the ability to assess immunopositivity, immunoprevalence, co-infections and incident infections (immunoconversions) to Helicobacter pylori, Campylobacter jejuni, Cryptosporidium parvum, Toxoplasma gondii, hepatitis A virus and noroviruses GI.I and GII.4 at several beaches throughout the US. Further, we’ve found evidence of asymptomatic norovirus and hepatitis A infections in visitors to a fecally contaminated beach. The assay produces results in as little as one hour and when used in conjunction with epidemiologic and water quality studies, provides valuable information that links human health effects more directly to water quality.
Phage display technology and molecular simulation combination to characterize anti-Opisthorchis viverrini antibodies
Authors: Siripanthong Sitthinon , Techasen Anchalee , Nantasenamat Chanin , Malik Aijaz Ahmad , Sithithaworn Paiboon , Leelayuwat Chanvit , Jumnainsong Amonrat 
Published: January 2023

Abstract
KKU505 hybridoma cell was used to produce monoclonal anti-Opisthrochis viverrini (OV) antibody. Unfortunately, hybridoma technology has many problems with instability for antibody production. Many studies have shown the possibility of developing a monoclonal antibody production using the antibody phage display. This paper presents antibody phage display technology for producing the phage display of the KKU505 Fab fragment and characterization using experiments and molecular simulations.
The characterization was successfully displayed on KKU505 Fab fragment phage particle using experimental means integrated with the molecular simulation. For the experiments, indirect ELISA was shown to be specific to the OV antigen in this KKU505 Fab fragment on the phage particle. Immunohistochemistry has been shown to have a localization and reaction pattern of phage presenting KKU505 Fab fragment with biliary mouse tissue infected with O. viverrini. In particular, the significance of the antibody-antigen dock was studied in order to gain a better understanding of the mechanisms underlying the KKU505 mAb and of the crude OV antigen interaction by Western blot.
All results showed that the fragment KKU505 Fab displayed on the phage particle has specific to the OV crude antigens. This study has shown that high potential phage displays technology shows monoclonal antibody production development. We have shown how the interactions of the KKU505 Fab fragment and OV antigen are evaluated in a successful application of experiment and molecular simulation. This fragment of the KKU505 Fab display on the particle is being developed to be used as a tool for diagnosing human opisthorchiasis.
Robust T- and B-cell clonotype detection using simultaneous targeted mRNA and immune repertoire profiling of single cells
Authors: Acob Ricelle A , Truong Tina , Levine Jay , Liu Kai , Jensen Devon , Prosen Dennis , Lazaruk Katherine 
Published: January 2023

Abstract
The key cell types contributing to the diversity of the immune repertoire are the T cells, with their T-cell receptor (TCR) and B cells with their B-cell receptor (BCR). During T- and B-cell development, the cell undergoes recombination of the germline gene segments, variable (V), diversity (D), and joining (J), also called V(D)J recombination. We developed a single cell clonotype detection assay for the BD Rhapsody™ Single-Cell Analysis System capable of processing thousands of single cells at once. We aimed to capture ~20,000 peripheral blood mononuclear cells from each individual donor (total of five) in a BD Rhapsody™ cartridge and took a sub-set of ~3,000 cells for immune repertoire profiling analysis of the complementarity determining region 3 (CDR3) of the variable regions of TCRα/TCRβ and the IgG heavy/light chains. The BD Rhapsody® Human Immune Response Panel for targeted mRNA gene expression analysis was used to classify the lymphocytes and monocyte populations. Results demonstrate that the BD Rhapsody™ VDJ assay can detect clonotype expression differences among different donors with &gt;80% sensitivity and specificity. The abundance of TCR and BCR transcripts was determined by the addition of a unique molecular index to each mRNA molecule. Overall, the current technology demonstrates the ability to evaluate immune populations using single cell next generation sequencing. This assay could be applied not only to study the immune repertoire but to understand the biology of immune-related diseases and immune response to therapies.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
BD, the BD Logo and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved.
Gentle cell sorting of immune cells for downstream single-cell RNA-seq
Authors: Jagnandan Nicole , Morachis Jose 
Published: January 2023

Abstract
Cell sorters have played an instrumental role in the progression of immunological discoveries. The ability to isolate specific immune cell types from a heterogenous population while maintaining high viability is critical to the discovery and generation of disease therapies. To isolate fragile and rare immune cell types, a gentle low pressure cell sorter is necessary. In this study, regulatory T cells (Tregs) were sorted from peripheral blood mononuclear cells using the WOLF® Cell Sorter developed by NanoCellect® and a traditional jet-in-air cell sorter. The isolated cells were then analyzed for single-cell RNA-seq using the 10x Genomics workflow. We also analyzed how different sorting methods affect the expression of stress markers induced during sorting. This study showed that despite the small percentage of Tregs in peripheral blood mononuclear cells, the WOLF Cell Sorter is able to obtain high purity, high viability, and high quality sequencing results. The WOLF Cell Sorter was also able analyze several subsets within the purified Treg populations in higher definition. Furthermore, fewer stress markers were induced when compared to the traditional jet-in-air cell sorter. This study provides novel insights into a robust and simplified workflow to use when isolating fragile immune cell types.
Label-free 3-D quantitative phase imaging cytometry with deep learning: identifying naive, memory, and senescent T cells
Authors: Ryu DongHun , Kim Kyunghwan , Cho Hayeon , Dao Khoi , Kim Young Seo , Ahn Daewoong , Min Hyun-Seok , Shin Eui-Cheol , Park YongKeun 
Published: January 2023

Abstract
The current prevailing methods for identifying immune cell subsets exploit a group of differentiation markers (CDs) targeted by fluorochrome or metal conjugated antibodies. However, such labeling methods, requiring a staining process and specific reagents, prevent rapid and cost-effective identification of immune cell subsets. Therefore we developed a label-free imaging cytometry platform that synergistically used refractive index (RI) tomography and three-dimensional (3-D) deep learning. We constructed and trained a deep learning classifier that learns unique representations from the 3-D RI map of each cell obtained using RI tomography without labeling. In this study, we were able to classify human naïve, memory, and senescent T cells according to the expression of CD4, CD8, CD45RA, CCR7 and CD57 using the label-free classifier within milliseconds, with high precision (&gt;95%) even though the morphological and biochemical characteristics extracted from the RI tomograms of the T cells are almost homogeneous. This cannot be achieved by conventional machine learning approaches that only exploit the set of manually extracted features. Our label-free cell sorting platform will facilitate rapid and cost-effective immunological and biomedical studies by eliminating the laborious labeling process.
Adarza’s ZIVA multiplex protein detection platform demonstrates flexibility with custom targets for monitoring both inflammation and anti-drug antibody response
Authors: Schmuke Jon J , Pillers Michelle , Mueller Ernie , Cooper Matt , Carter Alun , Kabakibi Ayman 
Published: January 2023

Abstract
We have used Arrayed Imaging Reflectometry (AIR™), a label-free protein microarray technology that relies on the target binding-induced perturbation of an antireflective coating on the surface of a silicon chip, to create a multiplex antibody array. ZIVA is a new multiplex protein detection platform that leverages AIR™ technology. Understanding the immunogenicity of a therapeutic protein is critical to the development of bioactive stability and efficacy. ZIVA technology can be customized to monitor for multiple biomarker types including markers of inflammation as well as specific ADA targets. In this example, biosensors were custom printed to monitor for the presence of an anti-drug antibodies generated against a single chain antibody presented on the surface of engineered CAR-T cells. Samples that are positive for ADA response may also be evaluated for presence of neutralizing antibodies (NAB) by evaluating specific controls also included on the biosensors. With array capabilities from low to hundreds of plex combined with a hands-free, automation-friendly workflow enabled by the ZIVA instrument, the technology has been designed for the ultimate user-experience.
High-throughput multi-dimensional single antigen-specific T cell profiling
Authors: Jiang Ning , Ma Ke-Yue , Schonnesen Alexandra A , Xia Amanda , Fan Yubo 
Published: January 2023

Abstract
Recent advances in single cell analysis has made it possible to interrogate the transcriptome of single cells in high-throughput manner. However, multi-dimensional profiling of antigen-specific T cells at single cell level for hundreds of antigens has not been realized, which becomes a bottleneck to the understanding of these important cells in immunological diseases.
Here we describe a new technology, TetTCR-SeqHD, that extends our previously developed technology, TetTCR-Seq for Tetramer associated TCR sequencing, to additional two dimensions, single cell gene expression and phenotype. In addition, we expand the throughput to tens of thousands of single T cells. We achieved these by using DNA-barcoded pMHC tetramers and antibodies in combination with single cell gene expression analysis, thus, paralleling all read-outs using next-gen sequencing.
To validate the technology, we have used previously generated polyclonal T cells to 6 different antigens with known TCR sequences. This analysis showed that we could achieve less than 5% on false discovery rate. We then used TetTCR-SeqHD on a panel of type 1 diabetes related peptides and showed unexpected features of antigen-specific T cells that separate healthy individuals from type 1 diabetes patients. In addition, TCR transgenics T cells confirmed their antigen specificity.
In summary, TetTCR-SeqHD enables us to analyze antigen-specific T cells with an extraordinary depth and breadth, which enables discoveries directly in humans. TetTCR-SeqHD would be a useful technology for studying many different diseases where screening for T cells antigens and deep profiling of antigen-specific T cells are highly sought after.
Donor deubiquitinase OTUD1 deficiency attenuates the severity of aGVHD mice after allo-HSCT by suppressing T cell activation and expansion
Authors: Cheng Qiao , Liu Yin , Wang Dong , Zhang Wenli , Xu Yang , Wu Depei 
Published: January 2023

Abstract
Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an important means to treat malignant diseases currently. However, the occurrence and development of acute graft versus host disease (aGVHD) seriously affect the survival and prognosis of patients. aGVHD is mainly mediated by the allogeneic immune response of activated donor T cells. Ubiquitination plays essential roles in T cell activation and is counteracted by multiple deubiquitinases (DUBs) with poorly understood. Here we show that OTUD1 deficiency negatively regulates the activation and effector function of naïve CD4+ T cells in vitro, and the proliferation of CD4+ T cells was also suppressed. Moreover, we found that OTUD1-deficient donor CD4+ T cells proliferates less in recipients compared to OTUD1+/+CD4+ T cells transferred. Interestingly, we found that OTUD1 expression was up-regulated in allo-HSCT mice when compared with syngeneic hematopoietic stem cell transplantation (syn-HSCT) mice. We further documented donor OTUD1 deficiency significantly prolongs the survival of aGVHD mice, and decreases the aGVHD scores. Moreover, OTUD1-deficient donor CD4+ T cells also attenuate the severity of aGVHD mice. Associated mechanistic studies revealed that the frequency and activation of CD4+ T cells were significantly reduced in recipients receiving OTUD1−/− bone marrow (BM) cells and splenocytes compared to those transferred OTUD1+/+ BM cells and splenocytes. Furthermore, overexpression of OTUD1 promotes NFAT-luciferase activity and up-regulates CD3-TCR complex stability. These results yield that targeting OTUD1 to regulate T cell-mediated allogeneic responses may be an effective therapeutic strategy to prevent and control aGVHD.
The role of TCF-1 in graft-versus-host disease and antitumor effects
Authors: Harris Rebecca , Karimi Mobin 
Published: January 2023

Abstract
Allogeneic hematopoietic stem cell transplants (HSCT) are used as a curative therapy for malignancies such as cancer. Residual tumor cells and recurrence are controlled by the graft-versus-tumor (GVT) effect, mediated by alloreactive T cells. However, these same cells also cause graft-versus-host disease (GVHD), a serious side effect. Naïve mature T cells in the graft recognize foreign HLA molecules on the host’s cells, and are alloactivated. The T cells then proliferate, produce cytokines, and migrate to the GVHD target organs (liver, skin, and small intestine), damaging host tissues. Separation of detrimental GVHD from beneficial GVT remains a serious unaddressed problem. Thus, the goal of this project is to achieve this separation through T cell signaling modulation.
The transcription factors T-bet and Eomes impact GVHD and GVT. They are downstream of T Cell Factor-1 (TCF-1), which is important in thymocytes and T cell activation. To determine the role of TCF-1 in alloactivation, mice with T cell-specific deletion of TCF-1 were phenotyped. These mice had increased expression of Eomes, Tbet, CD8+ central memory T cells, and polarization of CD44 expression. They also showed reduced activating T cells, and reduced expression of CXCR5 (a major chemokine receptor) when alloactivated. Finally, GVHD severity was reduced in mice transplanted with TCF-1-deficient T cells, and survival was improved in these mice. These data suggest that loss of TCF-1 results in a more activated state, yet GVHD is still reduced. Therefore, these cells may still be capable of antitumor immunity, separating GVHD from GVT. This work will help to improve outcomes for HSCT patients and enhance understanding of how mature alloactivated T cells are regulated.
Effect of CD3/CD28 activator on the cytotoxicity of human ex-vivo expanded γδ T cells against osteosarcoma cells
Authors: KO YUNMI , Lee Jun Ah 
Published: January 2023

Abstract

Background
We investigated the effect of CD3/CD28 activator on the ex-vivo expansion of human γδ T cells and their cytotoxic effect against osteosarcoma cells.

Methods
Peripheral blood mononuclear cells from healthy donors were culture in three different conditions for 10 days: basal expansion media (COM media), COM media with zoledronate+hrIL-2 (EX media), and EX media with CD3/CD28 activator (EX28 media). Percentage of expanded γδ T cells was analyzed by flow cytometry. IFN-γ and TNF-α secreted in the culture media was measured using ELISA assay. Cytotoxicity of γδ T cells against KHOS/NP cells was measured by MTT assay.

Results
Compared with COM media, the fold increase of cell expansion were 33.6 in EX media and 368.2 in EX28 media. The population of γδ T cells expanded in COM media represented 4.9% of CD3+ T cells, while γδ T cells expanded in EX media reached 71% of CD3+ T cells. The population of γδ T cells expanded in EX28 media represented 18% of CD25+ T cells, but not CD3+ T cells. The concentration of IFN-γ and TNF-α in the culture media increased both in EX (1000-fold, 340-fold) and EX28 (2340-fold, 1849-fold) media, compared with unstimulated control. Unexpectedly, IFN-γ and TNF-α production per cells were higher in EX media than in the EX28 media. The γδ T cells expanded in EX media showed potent cytotoxicity against KHOS/NP osteosarcoma cells in a ratio-dependent manner, but γδ T cells expanded in EX28 media did not show significant cytotoxic effect.

Conclusion
CD3/CD28 activator were not as potent as the combination of zoledronate and hrIL-2, for expansion γδ T cells nor its cytotoxic activity against KHOS/NP cells, although it increased the total number of T cells.
Combination therapy to enhance NK cell anti-tumor responses
Authors: Wolf Natalie K , Nicolai Chris , Dang Susanna , Synder Gail , Blaj Cristina , McWhirter Sarah , Picton Lora , Garcia K. Christopher , Raulet David H 
Published: January 2023

Abstract
Current cancer immunotherapies are targeted at mobilizing CD8 T cells, although numerous tumors have few or no antigens for CD8 T cells, or have lost MHC I, thereby evading CD8 T cells. There is therefore a need for improved therapies that mobilize other immune cells, such as NK cells, to the tumor microenvironment. A recent therapy candidate, cyclic dinucleotide (CDN), activates the cGAS-STING pathway of the innate immune system. Intratumoral CDN injections induce type I IFNs and other mediators that amplify the CD8 T cell response and induce regressions of primary tumors. MHC I-deficient tumors are not susceptible to CD8 T cell-mediated rejection, but data from our lab demonstrates that STING activation still leads to long-term tumor regressions, mediated by NK cells and in some cases CD4 T cells, in 30–100% of animals in 5/6 transplant models tested. In order to improve upon CDN monotherapy, we are testing combinations of CDN therapy with other therapies to target and activate NK cells in the tumor microenvironment and prevent or delay the onset of desensitization. Our results show that engineered IL-2 family cytokines synergize with CDN therapy to mobilize anti-tumor responses by NK cells and in some cases CD4 T cells. These anti-tumor responses are due to increased recruitment of NK cells to the tumor and sustained activation of these recruited NK cells. Our combination therapy regimen shows enhanced systemic effects, both in vivo in abscopal tumor models, and ex vivo by cytolysis of target cells mediated by splenocytes. Overall, our work demonstrates the impact of a novel combination therapy in mobilizing powerful NK cell-mediated anti-tumor activity, providing evidence for the potential of NK cell-targeted immunotherapeutics for cancer.
CD122-selective IL-2 complexes target γδ T and NK cells to reduce tumor-promoting Th17 effects and synergize with αPD-L1 to treat primary and metastatic bladder cancer
Authors: Reyes Ryan Michael , Deng Yilun , Zhang Deyi , Mukherjee Neelam , Ji Niannian , Wheeler Karen , Gupta Harshita B. , Garcia Myrna , Kornepati Anand , Svatek Robert S , Curiel Tyler J 
Published: January 2023

Abstract
αPD-L1 bladder cancer (BC) immunotherapy is effective in &lt;30% of cases. To address the large αPD-L1-unresponsive subset of patients, we tested αIL-2/IL-2 complexes (IL-2c) that block IL-2 from binding high-affinity IL-2Rα (CD25) for preferential IL-2Rβ (CD122) binding. Regulatory T cells (Tregs) capture IL-2 by CD25 whereas CD8+T, γδ T, and NK cells use CD122. We hypothesized that the tumor microenvironment, including local immune cells in primary versus metastatic BC, differentially affects immunotherapy responses. We used PD-L1+ mouse BC cell lines MB49 and MBT-2, for intravesical ([IVe] in bladder) and intravenous (IV) challenge studies of local versus metastatic BC. αPD-L1 or IL-2c alone reduced tumor burden and extended survival in IVe MB49 and MBT-2. Treg depletion using FOXP3DTR mice further enhanced IVe IL-2c effects, consistent with the known tumor-promoting role of Tregs in human BC. Using in vivo cell depletion approaches, we found that γδ T cells and NK cells, but not CD8+ T cells, were necessary for IL-2c efficacy in bladder. γδ T cells also reduced intratumoral Th17 cells that promote MB49 growth and are elevated in human BC. We confirmed γδ T cell effects in δ TCR KO mice, which abrogated IL-2c efficacy but not αPD-L1 efficacy. Neither αPD-L1 nor IL-2c alone treated metastatic MB49 and MBT-2 BC but the combination improved survival in both. These data are consistent with our recent findings in human BC patients in whom γδ T cell and NK cell cytotoxicity improved BCG immunotherapy. Thus, IL-2c is a promising novel BC immunotherapy that can improve bladder-specific immunity in primary BC. In metastatic BC, combination with αPD-L1 may also be a successful BC treatment strategy due to engagement of innate and adaptive immune responses.