About: Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

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An Entity of Type : schema:ScholarlyArticle, within Data Space : covidontheweb.inria.fr associated with source document(s)

Attributes	Values
type	Academic Article research paper schema:ScholarlyArticle
isDefinedBy	Covid-on-the-Web dataset
has title	Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells
Creator	Cheung, Anthony Correa, Isabel Crescioli, Silvia Figini, Mariangela Karagiannis, Panagiotis Lombardi, Sara Fink, Katja Lacy, Katie Spicer, James Ilieva, Kristina Karagiannis, Sophia Nestle, Frank Tutt, Andrew Luciani, Fabio Putterman, Chaim
Source	Medline; PMC
abstract	Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.
has issue date	2018-03-23(xsd:dateTime)
bibo:doi	10.3389/fimmu.2018.00493
bibo:pmid	29628923
has license	cc-by
sha1sum (hex)	d1232afc1f36390f6834e4627d99db439020be56
schema:url	https://doi.org/10.3389/fimmu.2018.00493
resource representing a document's title	Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells
has PubMed Central identifier	PMC5876289
has PubMed identifier	29628923
schema:publication	Front Immunol
resource representing a document's body	covid:d1232afc1f36390f6834e4627d99db439020be56#body_text
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