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Journal of Immunological Methods

Bharat Thyagarajan, Helene Barcelo, Eileen Crimmins, David Weir, Sharon Minnerath, Sithara Vivek, Jessica Faul
Variability induced by delayed cell processing and cell cryopreservation presents unique challenges for immunophenotyping in large population studies. We conducted a pilot study to evaluate the effect of delayed cell processing and cryopreservation on cell percentages obtained by immunophenotyping. We collected blood from 20 volunteers and compared the effect of (a) delayed cell processing up to 72 h (b) cryopreservation and (c) the combined effect of delayed cell processing and cryopreservation on immunophenotyping of 31 cell subsets that included several subsets of T, B, Natural Killer (NK) cells, monocytes and dendritic cells using both whole blood collected in EDTA tubes and peripheral blood mononuclear cells collected in CPT tubes...
September 14, 2018: Journal of Immunological Methods
Rhonda M Brand, Nabanita Biswas, Aaron Siegel, Ashley Myerski, Jarret Engstrom, E Jeffrey Metter, Randall E Brand, Ross D Cranston, Ian McGowan
BACKGROUND: There are limited data on the immunological responsiveness of healthy intestinal tissue when it is cultured and stimulated ex vivo. Such an ex vivo model has the potential to be a valuable tool in understanding disease pathogenesis and as a preclinical tool for the assessment of candidate therapeutic agents used to treat inflammatory bowel disease (IBD). AIM: We undertook a comprehensive study to evaluate ex vivo immunological responses of intestinal tissue and isolated mucosal mononuclear cells (MMC) to a broad range of stimuli...
September 12, 2018: Journal of Immunological Methods
Guillaume Wacht, Anh Poirot, Anne-Laure Charles, Mirjana Radosavljevic, Béatrice Uring-Lambert, Frédéric de Blay, Bernard Geny, Seiamak Bahram, Cindy Barnig
Here we extensively describe a FACS-based protocol for isolating intact non-stained human eosinophils from peripheral blood; a stop forward from our recently published initial study. This method of purification could be accomplished in <3 h with only small volumes of whole blood necessary, even in healthy subjects generally exhibiting low levels of circulating eosinophils. Eosinophil activation during the isolation steps appeared to be minimal and this purification procedure yielded high quality RNA. Moreover, these FACS-isolated eosinophils had prolonged viability in culture and were suitable for further activation assays...
September 11, 2018: Journal of Immunological Methods
Chandika Gamage, Yomani Sarathkumara, Virajie Weerakkodi, Kanae Shiokawa, Kumiko Yoshimatsu, Jiro Arikawa, Nobuo Koizumi
Monoclonal antibody (MAb)-based ELISA was developed to detect leptospiral antigens from the plasma of febrile patients. The MAb reacts specifically with pathogenic leptospires and the assay possesses excellent diagnostic test parameters compared to PCR, indicating that this new ELISA is useful for the early diagnosis of leptospirosis with low operational cost.
September 11, 2018: Journal of Immunological Methods
Erinke van Grinsven, Pieter H C Leliefeld, Janesh Pillay, Corneli W van Aalst, Nienke Vrisekoop, Leo Koenderman
Neutrophil antibacterial capacity is measured in animal models and in vitro as an important indicator of neutrophil function. To be able to extrapolate their conclusions, in vitro experiments should mimic the in vivo situation. In vivo, antibacterial capacity depends on multiple steps of bacterial sensing, priming, chemotaxis, phagocytosis and intracellular killing. Therefore, we developed a simply executed assay that involves multiple steps in one assay. The neutrophils were incorporated into a three-dimensional matrix of fibrin fibers, in which they could freely migrate...
September 8, 2018: Journal of Immunological Methods
Khin Myo Aye, Eiji Nagayasu, Minami Baba, Ayako Yoshida, Yasuhiro Takashima, Haruhiko Maruyama
Development of reliable, quantitative technologies for serodiagnosis of Toxoplasma gondii infection remains desirable. The luciferase immunoprecipitation system (LIPS) is a relatively simple, highly sensitive, and rapid quantitative immunoassay. The major advantages of this assay over ELISA are a wider dynamic range, shorter overall assay time, and less sample volume. In this study, we aimed to use this method for the serodiagnosis of toxoplasmosis. Recombinant Toxoplasma antigens (dense granule antigens GRA6, GRA7, and GRA8 and bradyzoite antigen BAG1) fused with nanoluciferase (Nluc, a small luciferase enzyme) were expressed in Escherichia coli, purified, and tested in LIPS assays with sera from experimental mice infected with T...
September 8, 2018: Journal of Immunological Methods
Niels Vande Casteele, Manon E Wildenberg
No abstract text is available yet for this article.
September 7, 2018: Journal of Immunological Methods
Lisa Holthaus, Daniel Lamp, Anita Gavrisan, Virag Sharma, Anette-Gabriele Ziegler, Martin Jastroch, Ezio Bonifacio
Dimethyl sulfoxide (DMSO) is a polar organic solvent used in a wide range of biological applications. DMSO is routinely used as a cryoprotectant for long-term cell freezing as well as to dissolve peptides and drugs for immune cell functional assays. Here, human CD4+ T cell activation, cytokine production, proliferation, and metabolism were investigated after stimulation in the presence of 0.01% to 1%, DMSO, representing concentrations commonly used in vitro. Surface expression of the activation markers CD69, CD25 and CD154 after polyclonal activation of CD4+ T cells was inhibited by 0...
September 7, 2018: Journal of Immunological Methods
Theodore Williams, Harsha S Krovi, Laurie G Landry, Frances Crawford, Niyun Jin, Anita Hohenstein, Megan E DeNicola, Aaron W Michels, Howard W Davidson, Sally C Kent, Laurent Gapin, John W Kappler, Maki Nakayama
Immortalized T cells such as T cell hybridomas, transfectomas, and transductants are useful tools to study tri-molecular complexes consisting of peptide, MHC, and T cell receptor (TCR) molecules. These cells have been utilized for antigen discovery studies for decades due to simplicity and rapidness of growing cells. However, responsiveness to antigen stimulation is typically less sensitive compared to primary T cells, resulting in occasional false negative outcomes especially for TCRs having low affinity to a peptide-MHC complex (pMHC)...
August 28, 2018: Journal of Immunological Methods
Eduardo Egea, Dary Mendoza, Gloria Garavito, Stephanie Saavedra, Humberto Gómez, Marcos Sanjuan
Conjugation of avian IgY antibodies to nanosensors has been extensively explored for the diagnostics of virus and parasite infection, as well as for the detection of pharmaceutically and toxicologically relevant molecules. However, to date this strategy has only been minimally applied the detection of allergens. In this study, gold nanoparticles (GNPs) were conjugated to a polyvalent IgY antibodies raised against Dermatophagoides group I allergens. GNPs were synthesized by HAuCL4 reduction using 1% trisodium citrate, and characterized them by absorption spectroscopy and transmission electron microscopy (TEM)...
August 27, 2018: Journal of Immunological Methods
Aljawharah Alrubayyi, Alexandra Schuetz, Kerri G Lal, Surat Jongrakthaitae, Kristopher M Paolino, Julie A Ake, Merlin L Robb, Mark S de Souza, Nelson L Michael, Dominic Paquin-Proulx, Michael A Eller
Antibody effector functions such as antibody dependent cellular cytotoxicity (ADCC) and antibody dependent cellular phagocytosis (ADCP) are considered important immunologic parameters following results from the RV144 clinical trial where a reduced risk of infection was associated with non-neutralizing antibody against the V1/V2 region of HIV envelope. The rapid and fluorometric ADCC (RFADCC) assay has been widely used to measure ADCC, however, the mechanism behind the activity measured remains unclear. Here, we demonstrate that monocytes acquire the PKH26 dye used in the RFADCC assay and that the commonly used RFADCC readout correlates with phagocytosis...
August 24, 2018: Journal of Immunological Methods
Mengjun Wang, Jiabin Shen, Harmin Herrera, Amit Singal, Charles Swindell, Lu Renquan, Anand Mehta
Hepatocellular carcinoma (HCC) accounts for >700,000 deaths worldwide, largely related to poor rates of diagnosis. Our previous work identified glycoproteins with increased levels of fucosylation in HCC. Plate-based assays to measure this change were compromised by increased levels of heterophilic antibodies with glycan lacking terminal galactose residues, which allowed for increased binding to the lectins used in these assays. To address this issue, we developed a multi-step protein A/G incubation and filtration method to remove the contaminating signal...
August 23, 2018: Journal of Immunological Methods
Di Xu, Xing Ming, Min Gan, Xiaoli Wu, Yang Dong, Dengyuan Wang, Hua Wei, Feng Xu
Cronobacter spp. (previously Enterobacter sakazakii), an opportunistic food-borne pathogen, involved in fatal infections in neonates and infants. Contaminated powdered infant formula (PIF) was considered to be the uppermost infective resource. In the present study, a thermophilic helicase-dependent isothermal amplification (tHDA) based method combined with silica-coated magnetic nanoparticles (Si-MNPs) as a DNA separator for rapid and sensitive detection of Cronobacter has been developed. The whole detection process can be completed in <3 h, with a detection limit of 100 and 101  CFU/ml Cronobacter in pure culture and artificially contaminated PIF, respectively, with a great sensitivity (94%) and specificity (100%)...
August 23, 2018: Journal of Immunological Methods
Cemile Sonmez, Figen Sezen, Selcuk Kilic
BACKGROUND: Serological methods have great importance for the detection of Treponema pallidum antibodies in syphilis diagnosis. The goal of the present study is to evaluate various commercially available screening assays in comparison with the FTA-abs test. METHODS: A total of 363 serum samples were enrolled in the study. Following routine testing including RPR and TPHA tests, each sample was tested by treponemal immunoassays (Chorus Syphilis Screen Recombinant, Architect Syphilis TP, Syphilis Virclia Monotest, Siemens Advia Centaur Syphilis, Euroimmun Treponema pallidum Screen ELISA, Vircell Syphilis ELISA IgG + IgM, SD Bioline Syphilis)...
August 20, 2018: Journal of Immunological Methods
Jung Seok Lee, Jaesung Kim, Se Pyeong Im, Si Won Kim, Jae Wook Jung, Jassy Mary S Lazarte, Jeong-Ho Lee, Kim D Thompson, Tae Sung Jung
Monomeric variable lymphocyte receptor B (VLRB) is one of the smallest binding scaffold (20-25 kDa) from jawless vertebrates, hagfish and lamprey. This relatively new class of binding scaffold has various advantages: i) it has a single peptide composition, amenable to molecular engineering for enhancing its stability and affinity; ii) it has a small size, contributing better tissue penetration and easier production using microorganism expression system. Monomeric arVLRB142, which can specifically bind to the glycoprotein of viral hemorrhagic septicemia virus (VHSV), was expressed in Pichia pastoris...
August 16, 2018: Journal of Immunological Methods
Amir A Sadighi Akha
The world is witnessing a rapid demographic shift towards an older population, a trend with major medical, social, economic and political implications. Aging is a multifaceted process, involving numerous molecular and cellular mechanisms in the context of different organ systems. A crucial component of aging is a set of functional and structural alterations in the immune system that can manifest as a decreased ability to fight infection, diminished response to vaccination, increased incidence of cancer, higher prevalence of autoimmunity and constitutive low-grade inflammation, among others...
August 13, 2018: Journal of Immunological Methods
Stephanie N Kennedy, Brittany Wilhite, J Margaret Castellini, Lorrie D Rea, Thomas B Kuhn, Andrea Ferrante, Todd M O'Hara
Immunoglobulins (Ig) are proteins that preserve immune homeostasis and are quantified to infer changes to the acquired humoral immune response in mammals. Measuring Ig in non-model wildlife for immune surveillance often requires ingenuity, and rigorous standardization of methodologies to provide reliable results especially when lacking species-specific reagents. We modified and optimized existing ELISA methodology utilizing the binding properties of Staphylococcus-derived Protein A (PrtA) to immunoglobulin G (IgG)...
August 9, 2018: Journal of Immunological Methods
Weifeng Xu, Michael Sank, Jennifer Cummings, Stephen Carl, Marina Juhel, Carol Gleason, Robert Dodge, Binodh S DeSilva, Gerry Kolaitis, Renuka Pillutla
Biological therapeutics are foreign antigens and can potentially induce immune response resulting in the formation of anti-drug antibodies (ADA), which in turn may lead to a wide range of side effects. Neutralizing Ab (NAb) is a subset of ADA that can bind to the pharmacological activity regions of therapeutic to inhibit or complete neutralize its clinical efficacy. A cell-based functional NAb assay is preferred to characterize its neutralization activity. However, cell-based NAb assays are often vulnerable to drug interference, as well as interference from numerous serum factors, including but not limited to growth factors and disease-related cytokines...
August 9, 2018: Journal of Immunological Methods
Sergi Luque, Marc Lúcia, Elena Crespo, Marta Jarque, Josep M Grinyó, Oriol Bestard
Emerging evidence suggests that donor-reactive memory B cells (mBC) play a key role inducing antibody-mediated rejection (ABMR) after solid organ transplantation and show a broader antigen repertoire than plasma cells thus, being potentially present even in absence of donor-specific antibodies. Therefore, the development of novel immune assays capable of quantifying circulating donor-reactive mBC in organ transplantation is highly warranted. We developed a novel HLA-specific B-cell FluoroSpot assay capable of enumerating multiple HLA-specific Antibody Secreting Cells (ASC) originated from circulating mBC after in-vitro polyclonal activation...
July 31, 2018: Journal of Immunological Methods
Mira Alt, Jessica Falk, Anna Maria Eis-Hübinger, Barbara Kropff, Christian Sinzger, Adalbert Krawczyk
Infections with the herpes simplex virus (HSV) and the human cytomegalovirus (HCMV) can lead to life-threatening diseases, particularly in immunosuppressed patients. Furthermore, HSV infections at birth (herpes neonatorum) can result in a disseminated disease associated with a fatal multiorgan failure. Congenital HCMV infections can result in miscarriage, serious birth defects or developmental disabilities. Antibody-based interventions with hyperimmunoglobulins showed encouraging results in clinical studies, but clearly need to be improved...
July 27, 2018: Journal of Immunological Methods
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