CD21 Recombinant Monoclonal Antibody

What is CD21 and why is it significant in immunological research?

CD21, also known as complement receptor type 2 (CR2), complement C3d receptor, or Epstein-Barr virus receptor, is a transmembrane protein expressed on B cells, follicular dendritic cells, thymocytes, and a subset of peripheral T cells . It contains a small cytoplasmic domain, a transmembrane region, and an extracellular domain consisting of 15 tandem short consensus repeat sequences . CD21's significance in immunological research stems from its multifunctional roles: it binds complement fragments C3d, C3dg, and iC3b; serves as the primary receptor for Epstein-Barr virus; and forms a complex with CD19 and CD81 that functions as a co-receptor to the B-cell receptor (BCR) . These properties make CD21 a crucial molecule for studying B cell activation, humoral immunity, and various disease states including systemic lupus erythematosus and EBV infection.

How do CD21 recombinant monoclonal antibodies differ from conventional CD21 antibodies?

CD21 recombinant monoclonal antibodies are produced using recombinant DNA technology, providing several distinct advantages over conventional monoclonal antibodies. Unlike hybridoma-derived antibodies, recombinant antibodies offer superior batch-to-batch consistency, higher specificity, and reduced background reactivity . The recombinant production process allows for precise genetic manipulation of the antibody sequence, enabling optimization of binding properties, affinity, and stability. Both mouse-derived (IgG1) and rabbit-derived recombinant monoclonal antibodies against CD21 are available, with the rabbit versions often providing higher sensitivity for certain applications . These advances in antibody engineering make recombinant CD21 antibodies particularly valuable for quantitative research applications where reproducibility and precision are paramount.

What is the molecular structure and functional domains of the CD21 protein that recombinant antibodies target?

CD21 is a 119 kDa (1092 amino acids) transmembrane glycoprotein with several distinct functional domains . Its structure includes:

Recombinant monoclonal antibodies can be engineered to target specific epitopes within these domains . For example, the Abcam antibody [CR2/1953] targets amino acids 1-200 within the extracellular domain . Different epitope specificities affect the antibody's capacity to recognize CD21 in various experimental contexts (native vs. denatured states, membrane-bound vs. soluble forms), which is crucial to consider when selecting antibodies for specific research applications.

What are the optimal applications for CD21 recombinant monoclonal antibodies in immune cell phenotyping?

CD21 recombinant monoclonal antibodies excel in immune cell phenotyping through several applications, with flow cytometry being the most widely utilized. For optimal flow cytometric analysis of CD21 expression:

• Use fresh or properly cryopreserved peripheral blood mononuclear cells (PBMCs)

• Co-stain with B cell markers (e.g., CD19) to identify CD21-expressing B cell subsets

• Titrate the recombinant antibody to determine optimal concentration (typically starting at 2-5 μg/mL)

Flow cytometric analysis can reveal distinct patterns of CD21 expression across B cell developmental stages and in disease states. For example, reduced CD21 expression identifies an anergic-like B cell population in autoimmune conditions. Additionally, CD21 recombinant antibodies can be employed in immunohistochemistry (paraffin sections) to visualize CD21+ follicular dendritic cell networks in lymphoid tissues and immunocytochemistry to study receptor distribution at the cellular level . The high specificity of recombinant antibodies makes them particularly valuable for multiplex phenotyping panels where cross-reactivity must be minimized.

How should researchers validate CD21 recombinant monoclonal antibodies for their specific experimental systems?

Rigorous validation of CD21 recombinant monoclonal antibodies is essential for ensuring experimental reliability. A comprehensive validation protocol should include:

• Positive control testing: Verify antibody performance using cell types known to express CD21 (e.g., human PBMCs, specifically B cells)

• Negative control testing: Confirm specificity by testing on CD21-negative cell populations or using isotype controls matched to the antibody's host species and isotype

• Application-specific validation:

  • For flow cytometry: Verify staining pattern on CD19+ B cells

  • For Western blot: Confirm detection of the expected 145 kDa band under non-reducing conditions

  • For IHC/ICC: Evaluate staining patterns in tissues with known CD21 expression profiles

• Cross-platform concordance: Compare results across multiple detection methods (e.g., flow cytometry vs. Western blot)

• Antibody titration: Determine the optimal antibody concentration that maximizes specific signal while minimizing background

This systematic approach to validation ensures that the selected CD21 recombinant monoclonal antibody will provide reliable and reproducible results in your specific experimental system.

What are the critical considerations when designing Western blot experiments using CD21 recombinant monoclonal antibodies?

Western blot experiments using CD21 recombinant monoclonal antibodies require careful consideration of several critical factors:

Following these guidelines will significantly improve the reliability and interpretability of Western blot experiments targeting CD21.

How can CD21 recombinant monoclonal antibodies be utilized to investigate B cell receptor (BCR) signaling pathways?

CD21 recombinant monoclonal antibodies offer sophisticated tools for dissecting BCR signaling pathways through several methodological approaches:

These methodologies provide comprehensive insights into how CD21 modulates BCR signaling and B cell activation in both normal and pathological conditions.

What approaches can researchers use to study the role of CD21 in Epstein-Barr virus infection using recombinant antibodies?

Researchers can employ CD21 recombinant monoclonal antibodies to investigate Epstein-Barr virus (EBV) infection mechanisms through several sophisticated experimental approaches:

• Viral binding and entry studies:

  • Use fluorescently-labeled EBV particles in combination with flow cytometry to quantify virus binding to CD21+ cells

  • Pre-treat cells with titrated amounts of CD21 recombinant antibodies to block specific binding epitopes and determine critical interaction domains

  • Employ super-resolution microscopy with labeled CD21 antibodies to visualize virus-receptor clustering during entry

• Genetic association studies:

  • Analyze HLA-DQ β1 alleles associated with EBV infectivity and EBV gp42 binding in conjunction with CD21 expression patterns

  • Use CD21 antibodies to sort cell populations with varying receptor densities to correlate with infection susceptibility

• CD21-EBV interaction specificity:

  • Utilize site-directed mutagenesis of CD21 combined with antibody epitope mapping to identify critical binding residues

  • Compare binding affinities of different EBV strains to CD21 using competitive inhibition with recombinant antibodies

• Functional consequences:

  • Measure changes in CD21 expression levels pre- and post-EBV infection using quantitative flow cytometry

  • Investigate CD21 shedding (producing soluble CD21) during EBV infection and its potential role in viral immune evasion

These methodological approaches collectively provide insights into the molecular mechanisms underlying CD21's role as the primary EBV receptor, potentially revealing targets for therapeutic intervention in EBV-associated diseases.

How can researchers use CD21 recombinant monoclonal antibodies to investigate complement-mediated immune responses?

CD21 recombinant monoclonal antibodies provide powerful tools for investigating complement-mediated immune responses through several sophisticated methodological approaches:

• C3d-antigen complex studies:

  • Generate C3d-antigen fusion proteins and use CD21 antibodies to block binding sites

  • Quantify how CD21-C3d interactions enhance B cell responses to specific antigens

  • Measure activation thresholds with and without CD21 engagement

• Complement cascade modulation:

  • Use CD21 antibodies in combination with complement inhibitors to dissect the relative contributions of different complement pathways

  • Develop in vitro systems where CD21-expressing cells are exposed to complement-opsonized targets while monitoring cellular responses

• Advanced imaging techniques:

  • Employ confocal microscopy with CD21 antibodies to visualize receptor clustering upon C3d binding

  • Use FRET (Förster resonance energy transfer) to measure molecular proximity between CD21 and other receptors during complement-mediated activation

• Functional assays of complement-dependent immune enhancement:

  • Measure B cell proliferation, antibody production, and memory formation in response to complement-decorated antigens

  • Compare responses when CD21-C3d interactions are blocked by specific antibodies

These approaches allow researchers to elucidate the mechanistic details of how CD21 engagement by complement fragments enhances B cell responses to antigens, a process critical for effective humoral immunity and potentially dysregulated in autoimmune conditions.

How should researchers address unexpected patterns of CD21 expression when using recombinant monoclonal antibodies?

When researchers encounter unexpected CD21 expression patterns using recombinant monoclonal antibodies, a systematic troubleshooting approach is essential:

• Antibody validation check:

  • Confirm the antibody recognizes the intended epitope by testing on known positive controls (human PBMCs, specifically B cells)

  • Verify antibody performance hasn't degraded by comparing with previous lot data or fresh aliquots

• Technical considerations:

  • For flow cytometry: Review compensation settings, evaluate potential spectral overlap, and check for nonspecific binding

  • For IHC/ICC: Assess fixation effects, as over-fixation can mask epitopes; try different antigen retrieval methods

  • For Western blot: Compare reducing vs. non-reducing conditions; CD21 detection is recommended under non-reducing conditions only

• Biological variables:

  • Consider developmental or activation-dependent changes in CD21 expression

  • Investigate presence of alternative splice variants with different epitope availability

  • Evaluate potential receptor internalization or shedding of soluble CD21

• Experimental design modifications:

  • Test multiple anti-CD21 antibody clones targeting different epitopes

  • Implement orthogonal detection methods (e.g., mRNA analysis via qPCR)

  • Use genetic approaches (e.g., CRISPR-edited cells) as definitive controls

By systematically addressing these factors, researchers can determine whether unexpected CD21 expression patterns represent technical artifacts or biologically meaningful variations that warrant further investigation.

What are common sources of data inconsistency when using CD21 recombinant monoclonal antibodies, and how can they be resolved?

Researchers frequently encounter data inconsistencies when using CD21 recombinant monoclonal antibodies. Understanding common sources of variability and their resolutions is critical:

Implementing these resolution strategies systematically will significantly improve data consistency and reliability when working with CD21 recombinant monoclonal antibodies across different experimental platforms.

How should researchers interpret variations in CD21 detection across different tissue types and disease states?

Interpreting variations in CD21 detection across different tissue types and disease states requires careful consideration of both biological and technical factors:

• Cell type-specific expression patterns:

  • B cells: CD21 expression varies by maturation stage and activation status; mature B cells express high levels while plasma cells downregulate expression

  • Follicular dendritic cells (FDCs): Express highest CD21 density of any cell type; critical for distinguishing FDC networks in lymphoid tissues

  • T cell subsets: Only certain T cell populations express CD21; levels are typically lower than on B cells

• Disease-associated variations:

  • Autoimmune conditions: Systemic lupus erythematosus (SLE) is associated with genetic variations in CD21 and altered expression patterns

  • B cell malignancies: Different lymphoma subtypes show characteristic CD21 expression; useful for classification

  • Infectious diseases: EBV infection can modulate CD21 expression; important for viral pathogenesis studies

• Methodological interpretation framework:

  • Establish baseline expression in healthy controls for each tissue type

  • Use multiple detection methods for confirmation (flow cytometry, IHC, Western blot)

  • Implement quantitative analysis where possible (mean fluorescence intensity, digital image analysis)

  • Consider CD21 detection in context with other markers (e.g., CD19, CD81)

• Technical considerations for interpretation:

  • Different antibody clones may have varying affinities for CD21 in different tissue contexts

  • Tissue processing can affect epitope availability differently across tissue types

  • When comparing disease states, match samples for processing methods and analysis platforms

This comprehensive approach enables researchers to distinguish meaningful biological variations from technical artifacts when interpreting CD21 detection across diverse experimental contexts.

How can CD21 recombinant monoclonal antibodies be engineered for therapeutic applications in autoimmune diseases?

Engineering CD21 recombinant monoclonal antibodies for therapeutic applications in autoimmune diseases represents an emerging frontier with several promising methodological approaches:

• Epitope-specific targeting strategies:

  • Engineer antibodies targeting specific CD21 domains that modulate B cell hyperactivity without compromising protective immunity

  • Develop antibodies that selectively block C3d-binding regions while preserving other CD21 functions

  • Design bispecific antibodies that simultaneously target CD21 and inhibitory receptors to induce targeted B cell anergy

• Antibody format optimization:

  • Explore various fragment formats (Fab, F(ab')2, scFv) to achieve optimal tissue penetration and half-life

  • Investigate different IgG isotypes to engage specific effector functions or avoid them entirely

  • Engineer Fc modifications to enhance or reduce complement activation depending on therapeutic goals

• Targeted delivery approaches:

  • Conjugate CD21 antibodies with cytotoxic agents for selective depletion of pathogenic B cell subsets

  • Develop antibody-drug conjugates targeting CD21-high B cells implicated in autoimmune pathogenesis

  • Design nanoparticle-based delivery systems decorated with CD21-binding domains for targeted immunomodulation

• Combination therapy strategies:

  • Investigate synergistic effects of CD21-targeted therapies with existing immunomodulatory agents

  • Develop rational combination approaches targeting multiple aspects of the CD21/CD19/CD81 complex

• Advanced preclinical testing:

  • Utilize humanized mouse models expressing human CD21 for preclinical evaluation

  • Implement ex vivo testing on patient-derived B cells to predict therapeutic efficacy

These engineering approaches could lead to novel therapeutics for conditions like systemic lupus erythematosus type 9 (SLEB9), which has established genetic associations with CD21 variations .

What are the emerging applications of CD21 recombinant monoclonal antibodies in studying prion diseases?

CD21 recombinant monoclonal antibodies are emerging as valuable tools for investigating prion diseases through several innovative methodological approaches:

• Prion trafficking studies:

  • Use CD21 antibodies to track the role of this receptor in targeting prions to follicular dendritic cells (FDCs)

  • Develop blocking antibodies to prevent CD21-mediated prion capture and investigate effects on neuroinvasion kinetics

  • Employ high-resolution imaging with fluorescently-labeled CD21 antibodies to visualize prion-receptor interactions

• Mechanistic investigations:

  • Utilize CD21 domain-specific antibodies to identify critical regions involved in prion binding

  • Compare binding affinities of different prion strains using competitive inhibition assays with recombinant antibodies

  • Investigate whether CD21-mediated signaling affects prion conversion rates or cellular susceptibility

• Therapeutic exploration:

  • Develop CD21-targeted approaches to block peripheral prion accumulation before neuroinvasion

  • Test whether disrupting CD21-prion interactions can delay disease onset in experimental models

  • Investigate potential for CD21 antibodies as diagnostic tools for detecting prion-infected FDCs in lymphoid biopsies

• Comparative cell biology:

  • Use recombinant antibodies to compare CD21 expression and function across species with differential prion susceptibility

  • Correlate CD21 polymorphisms with prion disease progression rates

These research directions could significantly advance our understanding of how CD21 contributes to peripheral prion replication and neuroinvasion, potentially opening new avenues for therapeutic intervention in these currently untreatable neurodegenerative diseases .

How can multi-omics approaches be integrated with CD21 recombinant monoclonal antibody studies to advance B cell immunology?

Integrating multi-omics approaches with CD21 recombinant monoclonal antibody studies offers powerful new methodologies for advancing B cell immunology:

• Antibody-facilitated cell sorting for downstream analysis:

  • Use CD21 recombinant antibodies for high-purity isolation of B cell subpopulations defined by CD21 expression levels

  • Apply single-cell RNA sequencing to CD21-sorted populations to identify transcriptomic signatures associated with receptor expression patterns

  • Perform ATAC-seq on isolated populations to identify chromatin accessibility differences associated with CD21 expression

• Spatial multi-omics applications:

  • Employ CD21 antibodies in spatial transcriptomics experiments to map receptor expression in tissue contexts

  • Use multiplexed imaging approaches (e.g., CODEX, CyTOF imaging) with CD21 antibodies to visualize receptor distribution relative to other cellular markers

  • Correlate CD21 expression patterns with tissue proteomics data to identify microenvironmental factors influencing receptor regulation

• Functional genomics integration:

  • Combine CRISPR screens with CD21 antibody-based phenotyping to identify genetic regulators of receptor expression and function

  • Use CD21 antibodies to monitor receptor expression changes following targeted genetic perturbations

  • Correlate genetic variants in CD21 pathway components with functional outcomes assessed by antibody-based assays

• Dynamic systems analysis:

  • Monitor temporal changes in CD21 expression during B cell activation using time-course experiments with recombinant antibodies

  • Correlate expression dynamics with phosphoproteomics data to map signaling network evolution

  • Develop computational models integrating antibody-derived quantitative data with multi-omics datasets