Peripheral blood cultured mast cells: Phenotypic and functional outcomes of different culture protocols
CONCLUSION: Four weeks of culture with IL-6 are sufficient to generate sizeable numbers of human mast cells from blood progenitors, thereby enabling simultaneous exploration of allergen-specific sIgE/FcεRI cross-linking and non-specific activation via MRGPRX2.PMID:33647250 | DOI:10.1016/j.jim.2021.113003 (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - March 1, 2021 Category: Allergy & Immunology Authors: Jessy Elst Vito Sabato Marie-Line M van der Poorten Margaretha Faber Athina L Van Gasse Leander P De Puysseleyr Chris H Bridts Christel Mertens Michel Van Houdt Marcus Maurer Margo M Hagendorens Didier G Ebo Source Type: research

Peripheral blood cultured mast cells: Phenotypic and functional outcomes of different culture protocols
CONCLUSION: Four weeks of culture with IL-6 are sufficient to generate sizeable numbers of human mast cells from blood progenitors, thereby enabling simultaneous exploration of allergen-specific sIgE/FcεRI cross-linking and non-specific activation via MRGPRX2.PMID:33647250 | DOI:10.1016/j.jim.2021.113003 (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - March 1, 2021 Category: Allergy & Immunology Authors: Jessy Elst Vito Sabato Marie-Line M van der Poorten Margaretha Faber Athina L Van Gasse Leander P De Puysseleyr Chris H Bridts Christel Mertens Michel Van Houdt Marcus Maurer Margo M Hagendorens Didier G Ebo Source Type: research

Calibration-free concentration analysis for quantification of anti-drug specific antibodies in polyclonal positive control antibodies and in clinical samples
J Immunol Methods. 2021 Feb 25:113002. doi: 10.1016/j.jim.2021.113002. Online ahead of print.ABSTRACTHighly sensitive assays for anti-drug antibodies (ADAs) are both a regulatory requirement and requisite for proper evaluation of the effects of immunogenicity on clinical efficacy and safety. Determination of ADA assay sensitivity depends on positive control antibodies to represent naturally occurring or treatment-induced ADA responses. An accurate determination of the proportion of drug-specific antibodies in these polyclonal positive control batches is critical for correct evaluation of assay sensitivity. Target purificat...
Source: Journal of Immunological Methods - February 28, 2021 Category: Allergy & Immunology Authors: Christina Aniol-Nielsen Henrik Toft-Hansen Madeleine Dahlb äck Claus Henrik Nielsen Helene Solberg Source Type: research

Calibration-free concentration analysis for quantification of anti-drug specific antibodies in polyclonal positive control antibodies and in clinical samples
J Immunol Methods. 2021 Feb 25:113002. doi: 10.1016/j.jim.2021.113002. Online ahead of print.ABSTRACTHighly sensitive assays for anti-drug antibodies (ADAs) are both a regulatory requirement and requisite for proper evaluation of the effects of immunogenicity on clinical efficacy and safety. Determination of ADA assay sensitivity depends on positive control antibodies to represent naturally occurring or treatment-induced ADA responses. An accurate determination of the proportion of drug-specific antibodies in these polyclonal positive control batches is critical for correct evaluation of assay sensitivity. Target purificat...
Source: Journal of Immunological Methods - February 28, 2021 Category: Allergy & Immunology Authors: Christina Aniol-Nielsen Henrik Toft-Hansen Madeleine Dahlb äck Claus Henrik Nielsen Helene Solberg Source Type: research

Highlights of the 33rd annual scientific meeting of the Association of Medical Laboratory Immunologists (AMLI)
J Immunol Methods. 2021 Feb 21:112994. doi: 10.1016/j.jim.2021.112994. Online ahead of print.ABSTRACTThe annual meeting of the Association of Medical Laboratory Immunologists (AMLI) was convened virtually over the month of August. Prior to the emergence of the COVID-19 pandemic, AMLI's scientific committee had chosen the following topics as the focus of its 2020 meeting: Histocompatibility Testing and Transplant Immunology; Secondary Immunodeficiency and Immunotherapy Monitoring; ANA Update; and Emerging Infectious Diseases and New Algorithms for Testing. Given the central role of the discipline in the evaluation of the ho...
Source: Journal of Immunological Methods - February 24, 2021 Category: Allergy & Immunology Authors: Vijaya Knight Medhat Z Askar Evangelos Ntrivalas Sarada L Nandiwada Lisa K Peterson Anne E Tebo Kamran Kadkhoda John L Schmitz Stanley J Naides Melissa R Snyder Amir A Sadighi Akha Source Type: research

Highlights of the 33rd annual scientific meeting of the Association of Medical Laboratory Immunologists (AMLI)
J Immunol Methods. 2021 Feb 21:112994. doi: 10.1016/j.jim.2021.112994. Online ahead of print.ABSTRACTThe annual meeting of the Association of Medical Laboratory Immunologists (AMLI) was convened virtually over the month of August. Prior to the emergence of the COVID-19 pandemic, AMLI's scientific committee had chosen the following topics as the focus of its 2020 meeting: Histocompatibility Testing and Transplant Immunology; Secondary Immunodeficiency and Immunotherapy Monitoring; ANA Update; and Emerging Infectious Diseases and New Algorithms for Testing. Given the central role of the discipline in the evaluation of the ho...
Source: Journal of Immunological Methods - February 24, 2021 Category: Allergy & Immunology Authors: Vijaya Knight Medhat Z Askar Evangelos Ntrivalas Sarada L Nandiwada Lisa K Peterson Anne E Tebo Kamran Kadkhoda John L Schmitz Stanley J Naides Melissa R Snyder Amir A Sadighi Akha Source Type: research

Highlights of the 33rd annual scientific meeting of the Association of Medical Laboratory Immunologists (AMLI)
J Immunol Methods. 2021 Feb 21:112994. doi: 10.1016/j.jim.2021.112994. Online ahead of print.ABSTRACTThe annual meeting of the Association of Medical Laboratory Immunologists (AMLI) was convened virtually over the month of August. Prior to the emergence of the COVID-19 pandemic, AMLI's scientific committee had chosen the following topics as the focus of its 2020 meeting: Histocompatibility Testing and Transplant Immunology; Secondary Immunodeficiency and Immunotherapy Monitoring; ANA Update; and Emerging Infectious Diseases and New Algorithms for Testing. Given the central role of the discipline in the evaluation of the ho...
Source: Journal of Immunological Methods - February 24, 2021 Category: Allergy & Immunology Authors: Vijaya Knight Medhat Z Askar Evangelos Ntrivalas Sarada L Nandiwada Lisa K Peterson Anne E Tebo Kamran Kadkhoda John L Schmitz Stanley J Naides Melissa R Snyder Amir A Sadighi Akha Source Type: research

Highlights of the 33rd annual scientific meeting of the Association of Medical Laboratory Immunologists (AMLI)
J Immunol Methods. 2021 Feb 21:112994. doi: 10.1016/j.jim.2021.112994. Online ahead of print.ABSTRACTThe annual meeting of the Association of Medical Laboratory Immunologists (AMLI) was convened virtually over the month of August. Prior to the emergence of the COVID-19 pandemic, AMLI's scientific committee had chosen the following topics as the focus of its 2020 meeting: Histocompatibility Testing and Transplant Immunology; Secondary Immunodeficiency and Immunotherapy Monitoring; ANA Update; and Emerging Infectious Diseases and New Algorithms for Testing. Given the central role of the discipline in the evaluation of the ho...
Source: Journal of Immunological Methods - February 24, 2021 Category: Allergy & Immunology Authors: Vijaya Knight Medhat Z Askar Evangelos Ntrivalas Sarada L Nandiwada Lisa K Peterson Anne E Tebo Kamran Kadkhoda John L Schmitz Stanley J Naides Melissa R Snyder Amir A Sadighi Akha Source Type: research

Functional recombinant human complement C1q with different affinity tags
We report here the generation of additional recombinant C1q proteins, with a FLAG tag fused to the C-terminus of C1qA or C1qB chains, or to the N-terminus of the C1qC chain. Two other variants harboring a Myc or a 6-His tag at the C-terminal end of C1qC were also produced. We show that all C1q variants, except for the His-tagged protein, can be produced at comparable yields and are able to bind with similar affinities to either IgM, a ligand of the globular regions, or to the C1r2-C1s2 tetramer, and to trigger IgM-mediated serum complement activation. These new recombinant C1q variants provide additional tools to investiga...
Source: Journal of Immunological Methods - February 23, 2021 Category: Allergy & Immunology Authors: Isabelle Bally Sarah Ancelet Jean-Baptiste Reiser V éronique Rossi Christine Gaboriaud Nicole M Thielens Source Type: research

Establishment of bioluminescent imaging model using murine T cell lymphoma susceptive to NK cell-dependent immune-surveillance
J Immunol Methods. 2021 Feb 17;491:112993. doi: 10.1016/j.jim.2021.112993. Online ahead of print.ABSTRACTAlthough the importance of NK cells as immune effector cells in controlling growth and metastatic dissemination of tumor cells has been widely recognized, it is unclear whether NK cells in different organs similarly control tumor cell growth and metastasis. In the present study, we established a bioluminescent imaging model of mouse T cell lymphoma cells, which are highly susceptive to NK cell-dependent immune-surveillance, to monitor the dissemination of lymphoma cells using an in vivo imaging system. The use of this m...
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Toshiyuki Fujiwara Kiho Miyazato Kei Takahashi Yoshihiro Hayakawa Source Type: research

Interactions of human mesenchymal stromal cells with peripheral blood mononuclear cells in a Mitogenic proliferation assay
CONCLUSIONS: A bidirectional interaction between MSCs and PBMCs occurs during co-culture. High numbers of MSCs inhibit PHA-stimulated PBMC proliferation and the PBMC response to stimulation; low numbers of MSCs augment these responses. Low density MSCs are susceptible to attrition, apparently by PBMC-induced apoptosis. These results may have direct application when considering therapeutic dosing of patients; low MSC doses may have unintended detrimental consequences.PMID:33609532 | DOI:10.1016/j.jim.2021.113000 (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Maryanne C Herzig Barbara A Christy Robbie K Montgomery Christopher P Delavan Katherine J Jensen Sarah E Lovelace Carolina Cantu Christi L Salgado Andrew P Cap James A Bynum Source Type: research

Array-based measurements of aero-allergen-specific IgE correlate with skin-prick test reactivity in asthma regardless of specific IgG4 or total IgE measurements
J Immunol Methods. 2021 Feb 17:112999. doi: 10.1016/j.jim.2021.112999. Online ahead of print.ABSTRACTSkin prick testing (SPT) and measurement of serum allergen-specific IgE (sIgE) are used to investigate asthma and other allergic conditions. Measurement of serum total IgE (tIgE) and allergen-specific IgG4 (sIgG4) may also be useful. The aim was to ascertain the correlation between these serological parameters and SPT. Sera from 60 suspected asthmatic patients and 18 healthy controls were assayed for sIgE and sIgG4 reactivity against a panel of 70 SPT allergen preparations, and for tIgE. The patients were also assessed by s...
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Aljali Hamed Ian Todd Patrick J Tighe Richard J Powell Tim Harrison Lucy C Fairclough Source Type: research

Establishment of bioluminescent imaging model using murine T cell lymphoma susceptive to NK cell-dependent immune-surveillance
J Immunol Methods. 2021 Feb 17:112993. doi: 10.1016/j.jim.2021.112993. Online ahead of print.ABSTRACTAlthough the importance of NK cells as immune effector cells in controlling growth and metastatic dissemination of tumor cells has been widely recognized, it is unclear whether NK cells in different organs similarly control tumor cell growth and metastasis. In the present study, we established a bioluminescent imaging model of mouse T cell lymphoma cells, which are highly susceptive to NK cell-dependent immune-surveillance, to monitor the dissemination of lymphoma cells using an in vivo imaging system. The use of this model...
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Toshiyuki Fujiwara Kiho Miyazato Kei Takahashi Yoshihiro Hayakawa Source Type: research

Interactions of human mesenchymal stromal cells with peripheral blood mononuclear cells in a Mitogenic proliferation assay
CONCLUSIONS: A bidirectional interaction between MSCs and PBMCs occurs during co-culture. High numbers of MSCs inhibit PHA-stimulated PBMC proliferation and the PBMC response to stimulation; low numbers of MSCs augment these responses. Low density MSCs are susceptible to attrition, apparently by PBMC-induced apoptosis. These results may have direct application when considering therapeutic dosing of patients; low MSC doses may have unintended detrimental consequences.PMID:33609532 | DOI:10.1016/j.jim.2021.113000 (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Maryanne C Herzig Barbara A Christy Robbie K Montgomery Christopher P Delavan Katherine J Jensen Sarah E Lovelace Carolina Cantu Christi L Salgado Andrew P Cap James A Bynum Source Type: research

Array-based measurements of aero-allergen-specific IgE correlate with skin-prick test reactivity in asthma regardless of specific IgG4 or total IgE measurements
J Immunol Methods. 2021 Feb 17:112999. doi: 10.1016/j.jim.2021.112999. Online ahead of print.ABSTRACTSkin prick testing (SPT) and measurement of serum allergen-specific IgE (sIgE) are used to investigate asthma and other allergic conditions. Measurement of serum total IgE (tIgE) and allergen-specific IgG4 (sIgG4) may also be useful. The aim was to ascertain the correlation between these serological parameters and SPT. Sera from 60 suspected asthmatic patients and 18 healthy controls were assayed for sIgE and sIgG4 reactivity against a panel of 70 SPT allergen preparations, and for tIgE. The patients were also assessed by s...
Source: Journal of Immunological Methods - February 20, 2021 Category: Allergy & Immunology Authors: Aljali Hamed Ian Todd Patrick J Tighe Richard J Powell Tim Harrison Lucy C Fairclough Source Type: research

A chimeric HLA-A2: β2M:Ig fusion protein for the study of virus-specific CD8 < sup > + < /sup > T-cells
CONCLUSION: The chimeric HLA-A2:β2M:Ig fusion protein-based assays provided a sensitive tool that may be paramount to measure virus-specific CD8+ T-cell response in a range of viral infections of clinical relevance.PMID:33600818 | DOI:10.1016/j.jim.2021.112997 (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 18, 2021 Category: Allergy & Immunology Authors: Ágata Lopes Ribeiro Franklin Pereira Ara újo Julia Pereira Martins Alice Aparecida Louren ço Jing Huang Felipe Valen ça Pereira Luis Adan Flores Andrade Adriana Alves Oliveira Paim Fl ávio Guimarães da Fonseca Edel Figueiredo Barbosa Stancioli Olind Source Type: research

Attenuation of GARP expression on regulatory T cells by protein transport inhibitors
This study aims to deduce the optimal stimulation period and the impact of protein transport inhibitors (PTIs), commonly used in the detection of intracellular cytokines, on GARP detection. Through flow cytometric analysis we analyzed different cell culture conditions for optimal GARP expression on activated Tregs. Healthy donor PBMCs were stimulated with either Staphylococcal Enterotoxin B (SEB) or PMA/Ionomycin (PMA/Iono), in the presence and absence of PTIs monensin and/or brefeldin A (BFA) and GARP expression was assessed on CD4+ CD25+ FOXP3+ Tregs. The optimal stimulation period for the detection of GARP was highest a...
Source: Journal of Immunological Methods - February 18, 2021 Category: Allergy & Immunology Authors: Sara Anvari Kimberly Schuster Andrea Grimbergen Carla M Davis George Makedonas Source Type: research

Establishment of bioluminescent imaging model using murine T cell lymphoma susceptive to NK cell-dependent immune-surveillance.
Abstract Although the importance of NK cells as immune effector cells in controlling growth and metastatic dissemination of tumor cells has been widely recognized, it is unclear whether NK cells in different organs similarly control tumor cell growth and metastasis. In the present study, we established a bioluminescent imaging model of mouse T cell lymphoma cells, which are highly susceptive to NK cell-dependent immune-surveillance, to monitor the dissemination of lymphoma cells using an in vivo imaging system. The use of this model is expected to be a highly sensitive method to examine the role of NK cells in con...
Source: Journal of Immunological Methods - February 17, 2021 Category: Allergy & Immunology Authors: Fujiwara T, Miyazato K, Takahashi K, Hayakawa Y Tags: J Immunol Methods Source Type: research

Interactions of human mesenchymal stromal cells with peripheral blood mononuclear cells in a Mitogenic proliferation assay.
CONCLUSIONS: A bidirectional interaction between MSCs and PBMCs occurs during co-culture. High numbers of MSCs inhibit PHA-stimulated PBMC proliferation and the PBMC response to stimulation; low numbers of MSCs augment these responses. Low density MSCs are susceptible to attrition, apparently by PBMC-induced apoptosis. These results may have direct application when considering therapeutic dosing of patients; low MSC doses may have unintended detrimental consequences. PMID: 33609532 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 17, 2021 Category: Allergy & Immunology Authors: Herzig MC, Christy BA, Montgomery RK, Delavan CP, Jensen KJ, Lovelace SE, Cantu C, Salgado CL, Cap AP, Bynum JA Tags: J Immunol Methods Source Type: research

Array-based measurements of aero-allergen-specific IgE correlate with skin-prick test reactivity in asthma regardless of specific IgG4 or total IgE measurements.
Abstract Skin prick testing (SPT) and measurement of serum allergen-specific IgE (sIgE) are used to investigate asthma and other allergic conditions. Measurement of serum total IgE (tIgE) and allergen-specific IgG4 (sIgG4) may also be useful. The aim was to ascertain the correlation between these serological parameters and SPT. Sera from 60 suspected asthmatic patients and 18 healthy controls were assayed for sIgE and sIgG4 reactivity against a panel of 70 SPT allergen preparations, and for tIgE. The patients were also assessed by skin prick tests for reactivity to cat, dog, house dust mite and grass allergens. Ov...
Source: Journal of Immunological Methods - February 17, 2021 Category: Allergy & Immunology Authors: Hamed A, Todd I, Tighe PJ, Powell RJ, Harrison T, Fairclough LC Tags: J Immunol Methods Source Type: research

A chimeric HLA-A2: β2M:Ig fusion protein for the study of virus-specific CD8+ T-cells.
CONCLUSION: The chimeric HLA-A2:β2M:Ig fusion protein-based assays provided a sensitive tool that may be paramount to measure virus-specific CD8+ T-cell response in a range of viral infections of clinical relevance. PMID: 33600818 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 15, 2021 Category: Allergy & Immunology Authors: Ribeiro ÁL, Araújo FP, Martins JP, Lourenço AA, Huang J, Pereira FV, Andrade LAF, Paim AAO, da Fonseca FG, Stancioli EFB, Martins-Filho OA, Peruhype-Magalhães V, Tsuji M, Coelho-Dos-Reis JG Tags: J Immunol Methods Source Type: research

Attenuation of GARP expression on regulatory T cells by protein transport inhibitors.
This study aims to deduce the optimal stimulation period and the impact of protein transport inhibitors (PTIs), commonly used in the detection of intracellular cytokines, on GARP detection. Through flow cytometric analysis we analyzed different cell culture conditions for optimal GARP expression on activated Tregs. Healthy donor PBMCs were stimulated with either Staphylococcal Enterotoxin B (SEB) or PMA/Ionomycin (PMA/Iono), in the presence and absence of PTIs monensin and/or brefeldin A (BFA) and GARP expression was assessed on CD4+ CD25+ FOXP3+ Tregs. The optimal stimulation period for the detection of GARP was highest a...
Source: Journal of Immunological Methods - February 15, 2021 Category: Allergy & Immunology Authors: Anvari S, Schuster K, Grimbergen A, Davis CM, Makedonas G Tags: J Immunol Methods Source Type: research

Identification of tumor-associated antigens of lung cancer: SEREX combined with bioinformatics analysis.
Abstract The aim of this study is to identify novel tumor-associated antigens (TAAs) of lung cancer by using serological analysis of recombinant cDNA expression library (SEREX) and bioinformatics analysis as well as to explore their humoral immune response. SEREX and pathway enrichment analysis were used to immunoscreen TAAs of lung cancer and elaborate their function in biological pathways, respectively. Subsequently, the sera level of autoantibodies against the selected TAAs (TOP2A, TRIM37, HSP90AB1, EEF1G and TPP1) was detected by immunoserological analysis to explore the immune response of these antigens. The ...
Source: Journal of Immunological Methods - February 12, 2021 Category: Allergy & Immunology Authors: Wang Y, Wang P, Liu M, Zhang X, Si Q, Yang T, Ye H, Song C, Shi J, Wang K, Wang X, Zhang J, Dai L Tags: J Immunol Methods Source Type: research

A multi-colour confocal microscopy method for identifying and enumerating macrophage subtypes and adherent cells in the stromal vascular fraction of human adipose.
This study examines leukocytes present in lymphoedema (LE) adipose tissue (AT) by multi-colour confocal microscopy. LE AT, collected by liposuction surgery, was digested with collagenase to separate adipocytes from other tissue cells, comprising blood and lymphatic endothelial cells, fibroblasts, and all vessel- and tissue-resident leukocytes - the stromal vascular fraction (SVF). SVF cells were activated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, adding Brefeldin-A to prevent cytokine secretion during the final 4 hours. Cells were incubated with CD11b-FITC and CD40-APC (M1 MØ)' or CD206-APC (M2 M&Osl...
Source: Journal of Immunological Methods - February 12, 2021 Category: Allergy & Immunology Authors: Dey A, Ni Z, Johnson MS, Sedger LM Tags: J Immunol Methods Source Type: research

Parameters of immunoglobulin extraction from dried blood spot cards and immunoassays for detection of antibody response to pathogens including the novel SARS-CoV-2.
In this study using different sources including serum, and monoclonal antibodies we established parameters for antibody extraction from the filter cards to assess antibody reactivity against Helicobacter pylori, measles virus (MV) and the novel coronavirus SARS-CoV-2 antigens. We demonstrated that DBS and dried undiluted serum result in completely preserved antibody activity for immunoassays, including in virus neutralization assays against MV. Extraction efficiency was determined by IgG concentration measurements. The plaque-reduction neutralization titer 50% of dried human serum spots remained stable after more than 10-d...
Source: Journal of Immunological Methods - February 11, 2021 Category: Allergy & Immunology Authors: Iankov I, Viker K, Turgeon C, Matern D, Galanis E Tags: J Immunol Methods Source Type: research

A sensitive method to quantify HIV-1 antibodies in mucosal samples.
Abstract Human immunodeficiency virus (HIV) remains a significant public health issue. In recent years, passive immunization with broadly neutralizing antibodies (bNabs) is being considered as a potentially efficacious approach for fighting HIV. One candidate that holds great promise is represented by the CD4-binding site targeted bNab capable of neutralizing over 90% of circulating HIV strains, VRC01. VRC01 along with its variants and clonal relatives - VRC01-LS and VRC07-523LS are currently being evaluated as vaccines in a number of clinical trials for HIV treatment and prevention. While mucosal areas of the bod...
Source: Journal of Immunological Methods - February 11, 2021 Category: Allergy & Immunology Authors: Prabhakaran M, Narpala S, Andrews SF, Connell SO, Lin CL, Coates EE, Flach B, Ledgerwood JE, McDermott AB Tags: J Immunol Methods Source Type: research

Non-invasive image-based cytometry for high throughput NK cell cytolysis analysis.
Abstract Natural Killer (NK) cells are lymphocytes that are the first line of defense against malignantly transformed cells, virally infected cells and other stressed cell types. To study the cytolytic function of NK cells in vitro, a cytotoxicity assay is normally conducted against a target cancerous cell line. Current assay methods are typically performed in mixed 2D cocultures with destructive endpoints and low throughput, thereby limiting the scale, time-resolution, and relevance of the assay to in vivo conditions. Here, we evaluated a novel, non-invasive, quantitative image-based cytometry (qIBC) assay for de...
Source: Journal of Immunological Methods - February 9, 2021 Category: Allergy & Immunology Authors: Patel RS, Lucas J, Timmins LM, Mukundan S, Teryek M, Bhatt R, Beaulieu A, Parekkadan B Tags: J Immunol Methods Source Type: research

Flow cytometric analysis of cell lineage and immune activation markers using minimal amounts of human whole blood-Field method for remote settings.
Abstract Remote laboratory settings - such as those where studies on neglected tropical diseases are performed - often lack specialized equipment required for flow cytometric analysis of immune cell subsets, which complicates evaluations on a single cell level using peripheral blood. Our aim was to establish a method to use whole blood for phenotypic characterization of T-cells for specific markers including CD3, CD4, HLA-DR, CD38, CCR5, CD27, CD45RA, CD25, and FoxP3. This method uses 100 μL whole blood which is stained for extracellular markers, lysed, and cryopreserved at -20 °C at a field laboratory ...
Source: Journal of Immunological Methods - February 8, 2021 Category: Allergy & Immunology Authors: Horn S, Ahmed MIM, Geldmacher C, Marandu TF, Osei-Mensah J, Debrah A, Layland LE, Hoerauf A, Kroidl I Tags: J Immunol Methods Source Type: research

Production, characterization and in-vitro applications of single-domain antibody against thyroglobulin selected from novel T7 phage display library.
Abstract Single domain antibodies (SdAbs) have been deployed in various biomedical applications in the recent past. However, there are no reports of their use in the immunoradiometric assays (IRMA) for thyroglobulin (Tg). Tg is the precursor molecule for the biosynthesis of thyroid hormones: thyroxine and triiodothyronine, which are essential for the regulation of normal metabolism in all vertebrates. Patients with differentiated thyroid cancer (DTC) require periodic monitoring of their serum thyroglobulin levels, as it serves as a prognostic marker for DTC. Here, we report a methodology to produce SdAbs against h...
Source: Journal of Immunological Methods - February 6, 2021 Category: Allergy & Immunology Authors: Jothivel K, Ghorui SK, Gholve C, Jain B, Dhekale Y, Gupta GD, Damle A, Banerjee S, Rajan MGR, Kulkarni S Tags: J Immunol Methods Source Type: research

Detection of alloreactive T cells from cryopreserved human peripheral blood mononuclear cells.
Abstract Precise analyses of alloreactive T cell phenotype and function can inform both the nature and intensity of adaptive responses to transplant antigens. However, alloreactive T cells are sparse and difficult to detect, particularly in cryopreserved peripheral blood mononuclear cells (PBMCs) and from hypo-responsive individuals. An assay to identify and phenotype alloreactive cells would be particularly valuable, especially for multi-center clinical trials that often store frozen samples for batch analysis. Herein we demonstrate consistent and reproducible alloreactive T cell detection in cryopreserved PBMC f...
Source: Journal of Immunological Methods - February 5, 2021 Category: Allergy & Immunology Authors: Tanimine N, Burrell BE, Deng K, Rickert C, Lee KM, Feeney N, Pardo J, LeGuern C, Markmann JF Tags: J Immunol Methods Source Type: research

Concordance with established tests and reproducibility of results obtained with a new single antigen chip array for HLA antibody detection (HISTO SPOT ® HLA AB).
In conclusion, the HISTO SPOT® HLA AB test is a good alternative to be used in addition or instead of the Luminex tests in clinical labs. PMID: 33549571 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - February 4, 2021 Category: Allergy & Immunology Authors: Launhardt K, Lefevre V, Souplet V, Prantl L, Marget M, Hovoricova B, Wenda S, Olivier C Tags: J Immunol Methods Source Type: research

Comprehensive epitope mapping using polyclonally expanded human CD8 T cells and a two-step ELISpot assay for testing large peptide libraries.
Abstract The genetic diversity of circulating HIV-1 strains poses a major barrier to the design, development and evaluation of HIV-1 vaccines. The assessment of both vaccine- and natural infection-elicited T cell responses is commonly done with multivalent peptides that are designed to maximally capture the diversity of potential T cell epitopes (PTEs) observed in natural circulating sequences. However, depending on the sequence diversity of viral subtypes and number of the HIV immunogens under investigation, PTE estimates, including HLA-guided computational methods, can easily generate enormous peptide libraries....
Source: Journal of Immunological Methods - January 30, 2021 Category: Allergy & Immunology Authors: Michelo CM, Dalel JA, Hayes P, Fernandez N, Fiore-Gartland A, Kilembe W, Tang J, Streatfield C, Gilmour J, Hunter E Tags: J Immunol Methods Source Type: research

Human peripheral basophils extended phenotype shows a high expression of CD244 immuno-regulatory receptor.
CONCLUSIONS: Our study shows that human resting Basophils express IgE and IgG Fc receptors and check point receptor CD244 that could potentially play a role in their previously reported immunoregulatory activity in sensitization and even in tumor immune escape. PMID: 33493550 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - January 22, 2021 Category: Allergy & Immunology Authors: Berger AE, Durrieu C, Dzviga C, Perrot JL, Lambert C Tags: J Immunol Methods Source Type: research

Whole blood can be used as an alternative to isolated peripheral blood mononuclear cells to measure in vitro specific T-cell responses in human samples.
In conclusion, we demonstrated that WB and PBMC methods of sample collection led to statistically concordant ICS results, indicating that WB-ICS is a suitable alternative to PBMC-ICS to analyze clinical trial samples. PMID: 33493551 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - January 22, 2021 Category: Allergy & Immunology Authors: Moris P, Bellanger A, Ofori-Anyinam O, Jongert E, Rodriguez JY, Janssens M Tags: J Immunol Methods Source Type: research

Systematic approach in macrophage polarization experiments: Maintaining integrity and reproducibility using flow cytometry and sample preparation.
Abstract Resolution of inflammation is an important physiological process following infection or injury. When inflammation fails to resolve, it can cause chronic inflammation, which exacerbates a myriad of diseases. Current anti-inflammatory treatment options are often inadequate to resolve inflammation, and as such, a key goal for drug discovery is to find natural products and novel compounds that can target immune resolution processes. In order to efficiently discovery new therapies, immune cell lines are often used, in conjunction with flow cytometry, to quickly and inexpensively screen potential drugs for immu...
Source: Journal of Immunological Methods - January 19, 2021 Category: Allergy & Immunology Authors: McCall JR, Sausman K Tags: J Immunol Methods Source Type: research

Monocyte monolayer assay in pre-transfusion testing: A magic key in transfusing patients with recurrent bad cross-match due to alloimmunization.
Abstract BACKGROUND: The monocyte monolayer assay (MMA) is an in-vitro assay that can predict the outcome of blood transfusion of antigen positive units when serologically compatible blood is not available. MATERIALS AND METHODS: Fifty-four patients testing positive by the antibody screening test using gel agglutination were further examined by the alloantibody identification panel to determine alloantibody specificity. After determining and categorizing the antibodies, patients' samples were examined using the MMA to determine the clinical significance of the detected alloantibodies. We also tested 2 seeding...
Source: Journal of Immunological Methods - January 16, 2021 Category: Allergy & Immunology Authors: El-Sayed HAN, Abdollah MR, Raafat SN, Ragab D Tags: J Immunol Methods Source Type: research

Relationship between circulating tumor-associated autoantibodies and clinical outcomes in advanced-stage NSCLC patients receiving PD-1/-L1 directed immune checkpoint inhibition.
Abstract BACKGROUND: Durable tumor regressions are observed in subset of advanced-stage non-small cell lung cancer (NSCLC) patients receiving PD-1/-L1 targeted immune checkpoint inhibitors (or 'immunotherapy') alone or combined with chemotherapy. However, the majority of advanced NSCLC patients receiving these agents do not experience long-term disease control. Existing methods to identify patients most likely to clinical benefit from PD-1/-L1 immunotherapy have limitations, creating a need for improved methods to guide selection of this treatment strategy, particularly to identify patients likely to benefit from ...
Source: Journal of Immunological Methods - January 9, 2021 Category: Allergy & Immunology Authors: Tarhoni I, Wakefield CJ, Kollipara R, Fidler MJ, Batus M, Bonomi P, Borgia JA Tags: J Immunol Methods Source Type: research

Depletion of CD45RA+ T cells: Advantages and disadvantages of different purification methods.
Conclusions CD45RA-depletion from G-CSF mobilized PBSC is feasible as one- and two-step approach and results in sufficient reduction of CD45RA+ T cells as well as B cells, but also to a co-depletion of NK cells. However, by gaining two independent cell products, the two-step approach enables the highest clinical flexibility in regard to individual graft composition with precise dosage of stem cells and T cells. PMID: 33417916 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - January 5, 2021 Category: Allergy & Immunology Authors: Bremm M, Krastel T, Cappel C, Zimmermann O, Pfeffermann LM, Katzki V, Bonig H, Schäfer R, Rettinger E, Merker M, Bremm S, Schaefer K, Klingebiel T, Soerensen J, Bader P, Huenecke S Tags: J Immunol Methods Source Type: research

Impact of complement and difference of cell-based assay and ELISA in determination of neutralization capacity against mumps and measles virus.
ssy B Abstract Neutralizing antibodies against mumps and measles virus are considered a correlate of protection against these diseases. Measurement of neutralizing antibodies is mostly performed using plaque reduction neutralization assay or 50% cell culture infective dose (CCID50) neutralization assay, but there are attempts for measuring neutralizing antibodies using enzyme-linked immunosorbent assay (ELISA) which is simpler, but the literature data regarding its convenience are diverse. The role of complement and antibodies in neutralizing capacity of sera is not completely defined. Here, CCID50 neutralization ...
Source: Journal of Immunological Methods - January 4, 2021 Category: Allergy & Immunology Authors: Brgles M, Kurtović T, Balija ML, Hećimović A, Mušlin T, Halassy B Tags: J Immunol Methods Source Type: research

Development of recombinant NS1-NS3 antigen based indirect ELISA for detection of bluetongue antibodies in sheep.
In this study, the applicability of the fusion construct has been further evaluated and optimised for field applicability. The fusion construct used in an ELISA platform projected a relative diagnostic sensitivity and specificity of 98.1% and 95.5% respectively against a pre-established test panel. The rNS1-NS3 ELISA showed substantially good agreement with the commercial BTV antibody detection kit. Finally, the study brings together the diagnostic capability of two NSPs, which can be a handy tool for sero-surveillance of bluetongue. PMID: 33412173 [PubMed - as supplied by publisher] (Source: Journal of Immunological Methods)
Source: Journal of Immunological Methods - January 4, 2021 Category: Allergy & Immunology Authors: Mohanty NN, Hemadri D, Archana M, Shetty N, Subramanyam V, Biswas SK, Chanda MM, Shivachandra SB Tags: J Immunol Methods Source Type: research

Development and characterization of an indirect ELISA to detect SARS-CoV  - 2 spike protein-specific antibodies.
In this study, we developed a SARS-CoV-2 spike (S1) protein enzyme-linked immunosorbent assay (ELISA) to detect SARS-CoV-2-specific antibodies. The SARS-CoV-2 S1 ELISA was found to be specific [97.8% (95% CI, 96.7% - 98.5%)], reproducible and precise (intra-assay coefficient of variability (CV) 5.3%, inter-assay CV 7.9%). A standard curve and the interpolation of arbitrary ELISA units per milliliter served to reduce the variability between different tests and operators. Cross-reactivity to other human coronaviruses was addressed by using sera positive for MERS-CoV- and hCoV HKU1-specific antibodies. Monitoring antibody d...
Source: Journal of Immunological Methods - January 4, 2021 Category: Allergy & Immunology Authors: Krähling V, Halwe S, Rohde C, Becker D, Berghöfer S, Dahlke C, Eickmann M, Ercanoglu MS, Gieselmann L, Herwig A, Kupke A, Müller H, Neubauer-Rädel P, Klein F, Keller C, Becker S Tags: J Immunol Methods Source Type: research

Quantitative assessment of NF κB transcription factor activity.
Quantitative assessment of NFκB transcription factor activity. J Immunol Methods. 2020 Dec 31;:112954 Authors: Hunter TTJ, Fear D, Lavender P, Spencer J, Peakman M, Ibrahim MAA Abstract The Nuclear Factor Kappa B (NFκB) pathway is an important signalling pathway in the immune system. Single gene defects in the NFκB pathway are described in a number of immunodeficiency diseases. These conditions provide a unique opportunity to investigate the mechanisms of NFκB function and how genetic mutations that disrupt this function lead to human disease. Here we describe a robust method f...
Source: Journal of Immunological Methods - December 31, 2020 Category: Allergy & Immunology Authors: Hunter TTJ, Fear D, Lavender P, Spencer J, Peakman M, Ibrahim MAA Tags: J Immunol Methods Source Type: research

Multiplex bead binding assays using off-the-shelf components and common flow cytometers.
Abstract The ability to quantify protein-ligand interactions in an accurate and high-throughput manner is important in diverse areas of biology and medicine. Multiplex bead binding assays (MBBAs) are powerful methods that allows for simultaneous analysis of many protein-ligand interactions. Although there are a number of well-established MBBA platforms, there are few platforms suitable for research and development that offer rapid experimentation at low costs and without the need for specialized reagents or instruments dedicated for MBBA. Here, we describe a MBBA method that uses low-cost reagents and standard cyt...
Source: Journal of Immunological Methods - December 22, 2020 Category: Allergy & Immunology Authors: Hattori T, Koide A, Panchenko T, Romero LA, Teng KW, Corrado AD, Koide S Tags: J Immunol Methods Source Type: research

Validation of a monoclonal antibody directed against the human sphingosine 1-phosphate receptor type 1.
MH Abstract The sphingosine 1-phosphate receptor type 1 (S1PR1) has several important functions, including stabilizing endothelial barrier and maintaining lymphocyte circulation. These functions are critically dependent on the regulation of S1PR1 cell surface expression. Currently available antibodies against human S1PR1 are not able to pick up cell surface expression on living cells by flow cytometry due to intracellular epitopes or unspecific binding. Here we describe the generation of a mouse monoclonal antibody specific for the N-terminal region of human S1PR1. It has an immunoglobulin M (IgM) kappa isotype a...
Source: Journal of Immunological Methods - December 21, 2020 Category: Allergy & Immunology Authors: Thuy AV, Paul JJ, Weigel C, Ziegler AC, Guntinas-Lichius O, Gräler MH Tags: J Immunol Methods Source Type: research

Detergent wash improves vaccinated lymph node handling ex vivo.
Abstract Lymph nodes (LNs) are essential secondary immune organs where the adaptive immune response is generated against most infections and vaccines. We recently described the use of live ex vivo LN slices to study the dynamics of adaptive immunity. However, when working with reactive lymph nodes from vaccinated animals, the tissues frequently became dislodged from the supportive agarose matrix during slicing, leading to damage that prevented downstream analysis. Because reactive lymph nodes expand into the surrounding adipose tissue, we hypothesized that dislodging was a result of excess lipids on the collagen c...
Source: Journal of Immunological Methods - December 14, 2020 Category: Allergy & Immunology Authors: Ball AG, Belanger MC, Pompano RR Tags: J Immunol Methods Source Type: research

Graphene oxide-gold nanoparticle-aptamer complexed probe for detecting amyloid beta oligomer by ELISA-based immunoassay.
Abstract Highly sensitive and easy detection method for Alzheimer's disease (AD) with a suitable biomarker is mandatory for preventing the factors resulting from AD. This research reports a modified ELISA with graphene for the detection of AD biomarker amyloid beta (Aβ) oligomer. Gold nanoparticle (AuNP) conjugated aptamer was used as the capture probe and attached on ELISA-graphene oxide surface through the amine linker. Antibody was used as the detection molecule to reach the maximum detection of Aβ oligomer. Suitable level of APTMS (2%), size of AuNP (30 nm) and aptamer concentration (2 μM) wer...
Source: Journal of Immunological Methods - December 14, 2020 Category: Allergy & Immunology Authors: Zhao J, Chang W, Liu L, Xing X, Zhang C, Meng H, Gopinath SCB, Lakshmipriya T, Chen Y, Liu Y Tags: J Immunol Methods Source Type: research

Monitoring Ravulizumab effect on complement assays.
Abstract Ravulizumab is a new C5 inhibitor therapeutic monoclonal antibody with a longer half-life than eculizumab. Monitoring complete complement blockade by eculizumab has allowed personalized therapy in specific settings. Similar action is expected with ravulizumab. Ravulizumab has 4 different amino acids from eculizumab, which allow greater affinity for the FcRn immunoglobulin receptor and change the affinity of the molecule for C5. Here we investigate if clinical lab tests traditionally used to monitor complement blockade for eculizumab are appropriate for monitoring complement blockade caused by ravulizumab....
Source: Journal of Immunological Methods - December 12, 2020 Category: Allergy & Immunology Authors: Willrich MAV, Ladwig PM, Martinez MA, Sridharan MR, Go RS, Murray DL, Complement Alternative Pathway Thrombotic Microangiopathy (CAP-TMA) Disease Oriented Group at the Mayo Clinic Tags: J Immunol Methods Source Type: research

Development of a lateral flow immunochromatography test for the rapid detection of bovine tuberculosis.
Abstract Detection of specific antibodies would be a useful test strategy for bovine tuberculosis (bTB) as a complement to the single skin test. We developed a lateral flow immunochromatography (LFIC) test for rapid bTB detection based on the use of a conjugate of gold nanoparticles with a recombinant G protein. After evaluating 3 Mycobacterium bovis (MB) antigens: ESAT-6, CFP-10 and MPB83 for the control line, we selected MPB83 given it was the most specific. The performance of the test was analyzed with 820 bovine sera, 40 sera corresponding to healthy animals, 5 sera from animals infected with Mycobacterium avi...
Source: Journal of Immunological Methods - December 12, 2020 Category: Allergy & Immunology Authors: Alonso N, Griffa N, Moyano RD, Mon ML, Colombatti Olivieri MA, Barandiaran S, Vivot MM, Fiorini G, Canal AM, Santangelo MP, Singh M, Romano MI Tags: J Immunol Methods Source Type: research

Development of competitive inhibition ELISA as an effective potency test to analyze human rabies vaccines and assessment of the antigenic epitope of rabies glycoprotein.
Abstract The potency of all modern tissue culture human rabies vaccines is measured based on the National Institute of Health (NIH) potency test that is laborious, time-consuming, involves large test variations and requires sacrifice of large number of animals. To circumvent these limitations, several researchers and WHO expert working groups have discussed development of alternative in vitro methods to replace the NIH potency test. Although several immunochemical methods have been proposed to quantify rabies glycoprotein (G-protein) using multiple murine monoclonal antibodies, we report an In vitro competitive in...
Source: Journal of Immunological Methods - December 9, 2020 Category: Allergy & Immunology Authors: Soni D, Sahoo I, Mallya AD, Kamthe P, Sahai A, Goel SK, Kulkarni PS, Dhere RM Tags: J Immunol Methods Source Type: research

Immunogenicity evaluating of the SLNs-alginate conjugate against Pseudomonas aeruginosa.
Abstract P. aeruginosa is of particular importance due to its numerous pathogens and the spread of its multidrug-resistant strains around the world. Hence there is a need to develop an effective vaccine to prevent the diseases with P. aeruginosa. The aim of present study was to evaluate the immunogenicity of alginate (Alg) antigen in conjugation with SLN as a candidate for nanovaccine against P. aeruginosa in mouse model. Alginate is a weak immunogen, but the immune responses produced by alginate are effective in killing Pseudomonas bacteria. To increase the immunogenicity of alginate, SLN was used that is useful ...
Source: Journal of Immunological Methods - November 28, 2020 Category: Allergy & Immunology Authors: Afshari H, Maleki M, Hakimian M, Tanha RA, Salouti M Tags: J Immunol Methods Source Type: research