SRF5 Antibody

Frequently Asked Questions on IRF5 Antibody Research
(Note: Corrected from "SRF5" to "IRF5" based on validated literature)

Advanced Research Questions

• How to resolve contradictory IRF5 expression data from different antibodies?

  • Case study: In human spleen, ab124792 detected IRF5 predominantly in red pulp, while HPA046700 showed diffuse staining.

    • Root cause: Epitope accessibility differences (linear vs. conformational) and fixation artifacts.

    • Solution:

      1. Perform antigen retrieval optimization (e.g., pH-adjusted citrate buffer for IHC).

      2. Validate with orthogonal methods (e.g., qRT-PCR on laser-captured IRF5+ vs. IRF5- cells) .

• What advanced techniques improve IRF5 epitope characterization?

  • Combine AlphaFold2-predicted structures with competitive ELISA:

    • Synthesize overlapping peptides spanning IRF5’s N-terminal DNA-binding domain (residues 1-150) and C-terminal regulatory region (residues 300-500).

    • Test antibody binding inhibition with these peptides to map epitopes .

  • Example finding: Antibodies targeting the IRF5 nuclear localization signal (residues 400-410) fail in IHC due to epitope masking during fixation .

• How to design a cross-species IRF5 study addressing antibody limitations?

  • Challenge: Most antibodies lack cross-reactivity between human/mouse IRF5.

  • Workflow:

    1. Use CRISPR knock-in mice expressing humanized IRF5.

    2. Validate antibodies in parallel assays (e.g., ab124792 for human IRF5 IHC, clone 6D3 for mouse IRF5 IB) .

    3. Employ spatial transcriptomics to correlate protein and mRNA expression in discordant regions.

Data Contradiction Analysis Framework

When IRF5 antibody results conflict with literature:

Adapted from ref , which found >70% of commercial IRF5 antibodies failed specificity tests.

Key Takeaways: