Nomenclature of CD molecules from the Tenth Human Leucocyte Differentiation Antigen Workshop

Clark, Georgina J.; Stockinger, Hannes; Balderas, Robert; Zelm, Menno C. van; Zola, Heddy; Hart, Derek N. J.; Engel, Pablo

doi:10.1038/cti.2015.38

Cited by 37 publications

(30 citation statements)

References 26 publications

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“…HLDA and CD workshops show successful examples of coordinated and community-driven efforts to bring order to the chaos of cell surface molecules targeting antibodies (35,70). The structured sharing of validation data should be amenable to the aggregation of relevant validation information and should guide the user to select quality reagents.…”

Section: Discussionmentioning

confidence: 99%

“…The structured sharing of validation data should be amenable to the aggregation of relevant validation information and should guide the user to select quality reagents. HLDA and CD workshops show successful examples of coordinated and community-driven efforts to bring order to the chaos of cell surface molecules targeting antibodies (35,70). EuroMabNet has focused on immunohistochemistry reagents (9).…”

Section: Discussionmentioning

confidence: 99%

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Relevance of Antibody Validation for Flow Cytometry

2019

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Antibody reagents are the key components of multiparametric flow cytometry analysis. Their quality performance is an absolute requirement for reproducible flow cytometry experiments. While there is an enormous body of antibody reagents available, there is still a lack of consensus about which criteria should be evaluated to select antibody reagents with the proper performance, how to validate antibody reagents for flow cytometry, and how to interpret the validation results. The achievements of cytometry moved the field to a higher number of measured parameters, large data sets, and computational data analysis approaches. These advancements pose an increased demand for antibody reagent performance quality. This review summarizes the codevelopment of cytometry, antibody development, and validation strategies. It discusses the diverse issues of the specificity, cross-reactivity, epitope, titration, and reproducibility features of antibody reagents, and this review discusses the validation principles and methods that are currently available and those that are emerging. We argue that significant efforts should be invested by antibody users, developers, manufacturers, and publishers to increase the quality and reproducibility of published studies. More validation data should be presented by all stakeholders; however, the data should be presented in sufficient experimental detail to foster reproducibility, and community effort shall lead to the public availability of large data sets that can serve as a benchmark for antibody performance.Flow cytometry has developed into an indispensable technique in the research and clinical investigation of immune and hematologic systems, with increasing applications in other cell biology disciplines. There are three main pillars in the practical application of this technology, namely the instrumentation, the analytical methods for large data sets, and the reagents used to design biological experiments. Despite dramatic developments in instrumentation (polychromatic (1), mass (2,3), and spectral (4,5) cytometry) and the significant developments in data analysis techniques for high content analyses (6-8), the last pillar of this trio remains consistent across time in its importance to the application: the fluorescent antibody conjugate.Our and others' experiences indicate that nearly half of antibodies, sold by companies or described by academic groups, do not function for the recommended application. They present staining patterns that conflict with those reported in the literature, show unexpected cross-reactivity, or have even failed the most basic specificity tests (9-11). There is growing alarm about results that cannot be reproduced by other research groups, including data published in high-impact journals. Antibodies are believed to be, in part, responsible for inconsistent experimental results and the publication of inaccurate data in the scientific literature (12,13).During recent years, both industry and academic groups have increased their efforts to increase the quality of t...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Relevance of Antibody Validation for Flow Cytometry

2019

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“…CD markers (accessible as a group by entering CD markers in the Annotations field) encode a heterogeneous group of cell surface proteins. The Human Leucocyte Differentiation Antigen (HLDA) workshop has designated 408 molecules (some of which are grouped within a CD) as CD markers [34]. Based upon our assembly and analysis, we could establish 1:1 orthology for 357 porcine genes to those that compose HLDA version 10.…”

Section: Utilitymentioning

confidence: 99%

The porcine translational research database: a manually curated, genomics and proteomics-based research resource

et al. 2017

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BackgroundThe use of swine in biomedical research has increased dramatically in the last decade. Diverse genomic- and proteomic databases have been developed to facilitate research using human and rodent models. Current porcine gene databases, however, lack the robust annotation to study pig models that are relevant to human studies and for comparative evaluation with rodent models. Furthermore, they contain a significant number of errors due to their primary reliance on machine-based annotation. To address these deficiencies, a comprehensive literature-based survey was conducted to identify certain selected genes that have demonstrated function in humans, mice or pigs.ResultsThe process identified 13,054 candidate human, bovine, mouse or rat genes/proteins used to select potential porcine homologs by searching multiple online sources of porcine gene information. The data in the Porcine Translational Research Database ((http://www.ars.usda.gov/Services/docs.htm?docid=6065) is supported by >5800 references, and contains 65 data fields for each entry, including >9700 full length (5′ and 3′) unambiguous pig sequences, >2400 real time PCR assays and reactivity information on >1700 antibodies. It also contains gene and/or protein expression data for >2200 genes and identifies and corrects 8187 errors (gene duplications artifacts, mis-assemblies, mis-annotations, and incorrect species assignments) for 5337 porcine genes.ConclusionsThis database is the largest manually curated database for any single veterinary species and is unique among porcine gene databases in regard to linking gene expression to gene function, identifying related gene pathways, and connecting data with other porcine gene databases. This database provides the first comprehensive description of three major Super-families or functionally related groups of proteins (Cluster of Differentiation (CD) Marker genes, Solute Carrier Superfamily, ATP binding Cassette Superfamily), and a comparative description of porcine microRNAs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-017-4009-7) contains supplementary material, which is available to authorized users.

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“…The lack of suitable markers can, in part, be addressed through antibody screening although there is limited availability of high quality antibodies to surface accessible epitopes. While approximately 370 antigens are targeted by monoclonal antibodies to cluster of differentiation (CD) molecules [8], this represents only a fraction of all molecules present on the cell surface (i.e., the surfaceome) and CD molecule screening has had limited success for nonhematopoietic stem cells [9–11]. Alternative approaches have been employed to evaluate the cell surface protein landscape based on transcriptomics [12], and physical and affinity enrichment strategies coupled with proteomics [13, 14], although each has limitations.…”

Section: Introductionmentioning

confidence: 99%

Mass Spectrometry-Based Identification of Extracellular Domains of Cell Surface N-Glycoproteins: Defining the Accessible Surfaceome for Immunophenotyping Stem Cells and Their Derivatives

Fujinaka

Waas

Gundry

2017

Methods in Molecular Biology

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Human stem cells and their progeny are valuable for a variety of research applications and have the potential to revolutionize approaches to regenerative medicine. However, we currently have limited tools to permit live isolation of homogeneous populations of cells apt for mechanistic studies or cellular therapies. While these challenges can be overcome through the use of immunophenotyping based on accessible cell surface markers, the success of this process depends on the availability of reliable antibodies and well-characterized markers, which are lacking for most stem cell lineages. This chapter outlines an iterative process for the development of new cell surface marker barcodes for identifying and selecting stem cell derived progeny of specific cell types, subtypes, and maturation stages, where antibody-independent identification of cell surface proteins is achieved using a modern chemoproteomic approach to specifically identify N-glycoproteins localized to the cell surface. By taking advantage of a large repository of available cell surfaceome data, proteins that are unlikely to confer cell type specificity can be rapidly eliminated from consideration. Subsequently, targeted quantitation by mass spectrometry can be used to refine candidates of interest, and a bioinformatic visualization tool is key to mapping experimental data to candidate protein sequences for the purpose of epitope selection during the antibody development phase. Overall, the process of developing cell surface barcodes for immunophenotyping is iterative and can include multiple rounds of discovery, refinement, and validation depending on the phenotypic resolution required.

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Nomenclature of CD molecules from the Tenth Human Leucocyte Differentiation Antigen Workshop

Cited by 37 publications

References 26 publications

Relevance of Antibody Validation for Flow Cytometry

Relevance of Antibody Validation for Flow Cytometry

The porcine translational research database: a manually curated, genomics and proteomics-based research resource

Mass Spectrometry-Based Identification of Extracellular Domains of Cell Surface N-Glycoproteins: Defining the Accessible Surfaceome for Immunophenotyping Stem Cells and Their Derivatives

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