CD79A Antibody, HRP conjugated

What is CD79A and what is its functional significance in B-cell biology?

CD79A (also known as Igα, MB-1) is a 25 kDa transmembrane protein consisting of 226 amino acid residues that functions as a critical component of the B-cell antigen receptor complex (BCR). It cooperates with CD79B (Igβ) to initiate signal transduction cascades following antigen binding to the BCR. This interaction leads to complex internalization, trafficking to late endosomes, and subsequent antigen presentation . CD79A is also essential for BCR surface expression and efficient differentiation of pro- and pre-B-cells. At the molecular level, it stimulates SYK autophosphorylation and activation, binds to BLNK to facilitate SYK-mediated BLNK phosphorylation, and interacts with Src-family tyrosine kinases to enhance their activity . Interestingly, CD79A also serves a regulatory function by repressing BCR signaling during the development of immature B-cells, highlighting its multifaceted role in B-cell maturation and function .

What are the primary applications for CD79A Antibody, HRP conjugated in laboratory research?

CD79A Antibody, HRP conjugated has multiple validated applications in research settings:

• Western Blotting (WB): Useful for detecting CD79A protein in cell and tissue lysates at dilutions ranging from 1:300-5000 .

• Immunohistochemistry on paraffin-embedded tissues (IHC-P): Effective at dilutions of 1:200-400 for identifying CD79A-expressing cells in tissue sections .

• Immunohistochemistry on frozen tissues (IHC-F): Can be used at dilutions of 1:100-500 for detecting CD79A in frozen tissue specimens .

The HRP conjugation eliminates the need for secondary antibody incubation, streamlining experimental workflows and potentially reducing background signal in sensitive applications . These applications make this antibody particularly valuable for studying B-cell development, lymphoma classification, and immune response mechanisms in human samples.

How can researchers distinguish between normal and abnormal CD79A expression patterns?

Normal CD79A expression is characterized by consistent, membranous staining in B lymphocytes with moderate to strong intensity. In contrast, abnormal expression patterns have important diagnostic implications. In classic Hodgkin lymphoma (CHL), CD79A expression in Hodgkin Reed-Sternberg (HRS) cells typically shows weaker staining intensity than normal B-cells and plasmacytes, with homogeneous cytoplasmic distribution . Researchers should note that only a subset of CHL cases (approximately 36% based on one study) express CD79A in HRS cells, with varying proportions (10-100%) of positive cells .

When evaluating CD79A expression, it's crucial to assess:

• Staining intensity relative to normal B-cells

• Subcellular localization (membrane vs. cytoplasmic)

• Proportion of positive cells

• Homogeneity of staining across the cell population

Dual immunohistochemistry using CD79A with other markers (like CD30 for HRS cells) can help identify cell populations with aberrant phenotypes, as some HRS cells may be CD30+/CD79a+ while others are CD30+/CD79a- .

What are the optimal experimental conditions for using CD79A Antibody, HRP conjugated in Western blotting?

When performing Western blotting with CD79A Antibody, HRP conjugated, researchers should implement the following protocol for optimal results:

• Sample preparation: Prepare lysates from B-cell lines (such as Daudi, Raji, or Ramos Burkitt's lymphoma cell lines) using a compatible lysis buffer .

• Separation: Use reducing conditions for electrophoresis, as CD79A detection is validated under reducing conditions .

• Transfer: PVDF membrane is recommended based on validated protocols .

• Dilution: Use the antibody at 1:300-5000 dilution depending on sample type and expression level .

• Expected bands: Look for specific bands at approximately 44-52 kDa for CD79A .

• Controls: Include positive controls (B-cell lines) and negative controls (non-B-cell lines) to validate specificity.

For troubleshooting weak signals, researchers can adjust antibody concentration, extend incubation time, or optimize protein loading. When detecting low abundance of CD79A, enhanced chemiluminescence detection systems may improve sensitivity.

What are the clinicopathological implications of CD79A expression in lymphoma research?

CD79A expression analysis has significant clinicopathological value in lymphoma research, particularly in classic Hodgkin lymphoma (CHL). Studies have revealed that CD79A expression in HRS cells correlates with poorer clinical outcomes:

These findings suggest that CD79A expression status could serve as a valuable prognostic marker in CHL, potentially informing treatment decisions and follow-up strategies.

How should researchers design experiments to investigate CD79A signaling pathways?

When investigating CD79A signaling pathways, researchers should consider the following experimental design strategies:

• Stimulation models: Design experiments that trigger BCR activation using anti-IgM antibodies or specific antigens to observe CD79A phosphorylation and downstream signaling.

• Co-immunoprecipitation studies: Utilize CD79A antibodies for pull-down assays to identify interaction partners, particularly focusing on:

  • BLNK (SLP-65) interactions

  • SYK recruitment and activation

  • Interactions with Src-family tyrosine kinases

• Phosphorylation analysis: Employ phospho-specific antibodies to monitor the phosphorylation status of CD79A ITAM motifs and downstream effectors following BCR stimulation.

• Mutagenesis approaches: Generate CD79A mutants to identify critical residues required for:

  • BCR complex formation

  • Signal transduction

  • Endocytosis and trafficking

• Inhibitor studies: Use specific kinase inhibitors (SYK inhibitors, BTK inhibitors) to dissect the contribution of different signaling components to CD79A-mediated pathways.

When designing these experiments, researchers should include appropriate controls, such as CD79A-deficient cells or isotype control antibodies, to validate the specificity of observed effects.

What are the recommended antigen retrieval methods for CD79A immunohistochemistry?

For optimal CD79A detection in immunohistochemistry on paraffin-embedded tissues (IHC-P), heat-induced epitope retrieval (HIER) is strongly recommended. Based on validated protocols, researchers should implement the following approach:

• Retrieval solution: Use a basic antigen retrieval reagent (such as VisUCyte Antigen Retrieval Reagent-Basic or equivalent) .

• Heating method: Apply heat treatment prior to primary antibody incubation to unmask epitopes.

• Incubation conditions: After retrieval, apply CD79A antibody at 5 μg/mL (or at dilutions of 1:200-400) for 1 hour at room temperature .

• Detection system: For HRP-conjugated primary antibodies, proceed directly to chromogenic development. For unconjugated primary antibodies, use an appropriate HRP-polymer detection system .

• Counterstaining: Hematoxylin counterstaining provides good contrast to visualize tissue architecture.

This protocol has been validated for lymphoid tissues, where specific CD79A staining localizes to lymphocytes . For frozen sections (IHC-F), fixation in acetone or paraformaldehyde followed by permeabilization is recommended, with antibody dilutions in the range of 1:100-500 .

What controls should be included when using CD79A Antibody, HRP conjugated?

To ensure experimental validity and interpretation of results using CD79A Antibody, HRP conjugated, researchers should include the following controls:

For flow cytometry applications, including a viability dye is essential to exclude dead cells that may bind antibodies non-specifically. For Western blot applications, loading controls such as GAPDH or β-actin should be used to normalize protein loading across samples.

How can researchers accurately quantify CD79A expression in tissue samples?

Accurate quantification of CD79A expression in tissue samples requires standardized approaches and appropriate analysis methods:

• Digital image analysis for IHC:

  • Capture high-resolution images using consistent exposure settings

  • Use automated software to segment CD79A-positive cells

  • Quantify parameters including positive cell percentage, staining intensity, and H-score (combining intensity and percentage)

  • Compare to internal controls within the same section for relative quantification

• Scoring systems for manual evaluation:

  • Percentage scoring: Determine the percentage of CD79A-positive cells (e.g., 10-20%, 20-30%, etc.)

  • Intensity scoring: Grade staining intensity relative to normal B-cells (weaker, equal, stronger)

  • Combined scores: Calculate H-scores by multiplying intensity (0-3) by percentage (0-100%)

• Flow cytometry quantification:

  • Use mean fluorescence intensity (MFI) to assess expression levels

  • Calculate the ratio of sample MFI to isotype control MFI

  • Employ standardized beads for absolute quantification

• Western blot quantification:

  • Use densitometry software to measure band intensity

  • Normalize to housekeeping proteins

  • Include concentration standards for absolute quantification

For scientific publications, researchers should clearly describe the quantification methodology, including thresholds for positivity, scoring criteria, and statistical approaches used for comparative analyses.

How does CD79A antibody performance compare across different species models?

CD79A exhibits varying degrees of conservation across species, which impacts antibody reactivity and experimental design considerations when working with different model organisms:

When working with non-human models, researchers should:

• Verify sequence homology in the antibody's epitope region

• Conduct preliminary validation experiments

• Consider species-specific antibodies for critical experiments

• Include appropriate positive controls from the species being studied

The CD79A Monoclonal Antibody, HRP Conjugated from Bioss (catalog #bsm-60791r-hrp) has been reported to react with human samples, with potential cross-reactivity in other species requiring validation by individual laboratories .

What are the most effective strategies for multiplexed detection of CD79A with other B-cell markers?

Multiplexed detection of CD79A with other B-cell markers provides comprehensive characterization of B-cell populations and their states. The following strategies are recommended for effective multiplexed analysis:

• Sequential immunohistochemistry:

  • Perform CD79A staining first using HRP-conjugated antibody and DAB (brown) visualization

  • Follow with heat-mediated antibody stripping

  • Apply second primary antibody (e.g., CD20) with alkaline phosphatase detection and contrasting chromogen (e.g., Fast Red)

  • This approach has been validated for dual CD30/CD79A staining in Hodgkin lymphoma studies

• Multiplex immunofluorescence:

  • Utilize CD79A antibody with appropriate fluorophore-conjugated secondary antibody

  • Combine with antibodies against other B-cell markers (CD19, CD20, CD22)

  • Include lineage markers (CD3 for T-cells) for contextual analysis

  • Apply spectral unmixing to resolve overlapping fluorescence signals

• Multi-parameter flow cytometry:

  • Include CD79A in antibody panels with CD19, CD20, CD27 (memory B-cells), and CD38 (plasma cells)

  • Add functional markers like IgD, IgM, and activation markers

  • Use viability dyes and doublet discrimination for clean population isolation

• Single-cell sequencing with protein detection:

  • Combine CD79A antibody detection with single-cell RNA sequencing

  • Correlate protein expression with transcriptional signatures

  • Identify novel B-cell subpopulations based on combined profiles

When designing multiplexed panels, researchers should carefully select antibody clones to minimize cross-reactivity and validate the specificity of each marker in the multiplexed context.

How can researchers leverage CD79A expression analysis for lymphoma classification and prognosis?

CD79A expression analysis offers valuable insights for lymphoma classification and prognostication that can enhance diagnostic accuracy and treatment decision-making:

Researchers should integrate CD79A analysis with other established prognostic factors to develop comprehensive risk assessment models for lymphoma patients.

What are the current limitations in CD79A antibody technology and how might they be addressed?

Current CD79A antibody technology faces several limitations that impact research applications. Understanding these challenges and potential solutions can help researchers optimize experimental design:

• Epitope accessibility issues:

  • Challenge: CD79A's membrane localization and association with other BCR components can limit antibody access to epitopes

  • Solution: Develop antibodies targeting more accessible regions and optimize antigen retrieval protocols

• Post-translational modification detection:

  • Challenge: Current antibodies may not distinguish between phosphorylated and non-phosphorylated forms

  • Solution: Generate phospho-specific CD79A antibodies targeting key regulatory residues

• Isoform specificity:

  • Challenge: Limited ability to distinguish between the reported CD79A isoforms

  • Solution: Develop isoform-specific antibodies with validated epitope mapping

• Cross-reactivity concerns:

  • Challenge: Variable performance across species due to sequence differences

  • Solution: Generate species-specific antibodies with thoroughly validated cross-reactivity profiles

• Quantification standardization:

  • Challenge: Lack of universal standards for quantitative CD79A expression analysis

  • Solution: Develop calibrated reference materials and standardized reporting metrics

Future antibody technologies, including recombinant antibody engineering and nanobody development, may address these limitations and enhance the specificity, sensitivity, and reproducibility of CD79A detection across diverse research applications.

How can researchers integrate CD79A analysis with other emerging technologies in immunology research?

Integration of CD79A analysis with cutting-edge technologies creates opportunities for deeper insights into B-cell biology and lymphoma research:

• Spatial transcriptomics and CD79A protein expression:

  • Combine CD79A immunohistochemistry with spatial transcriptomics

  • Map CD79A protein expression relative to transcriptional signatures

  • Identify microenvironmental factors influencing CD79A expression

• Mass cytometry (CyTOF) applications:

  • Incorporate metal-tagged CD79A antibodies in high-parameter panels

  • Simultaneously analyze dozens of markers to characterize B-cell subsets

  • Correlate CD79A expression with signaling pathway activation

• CRISPR-Cas9 functional genomics:

  • Screen for genes that regulate CD79A expression or function

  • Generate CD79A reporter cell lines for high-throughput screening

  • Create CD79A mutant models to dissect functional domains

• Single-cell multi-omics:

  • Integrate CD79A protein detection with transcriptomics and epigenomics

  • Profile CD79A+ cells at single-cell resolution across multiple modalities

  • Identify regulatory networks controlling CD79A expression

• Machine learning analysis:

  • Apply convolutional neural networks to automate CD79A+ cell identification

  • Develop predictive models for patient outcomes based on CD79A expression patterns

  • Discover novel CD79A expression signatures associated with disease subtypes

These integrated approaches will enable researchers to move beyond descriptive studies toward mechanistic understanding of CD79A biology and its implications for B-cell function in health and disease.