HIST1H2BC (Ab-15) Antibody

What is HIST1H2BC and what is its biological significance?

HIST1H2BC (Histone H2B type 1-C/E/F/G/I) is a core histone protein that plays a critical role in packaging DNA into chromatin and regulating gene expression. It is essential for maintaining the structure and stability of the genome, making it a key player in cellular processes such as transcription, DNA replication, and DNA repair .

The protein is one of several histone H2B variants and is encoded by the HIST1H2BC gene. Understanding its function is crucial for unraveling the mechanisms of gene regulation and chromatin dynamics in both normal development and disease progression .

What are the key specifications of the HIST1H2BC (Ab-15) Antibody?

The HIST1H2BC (Ab-15) Antibody is a polyclonal antibody produced in rabbits that specifically targets the region around lysine 15 of the Human Histone H2B type 1-C/E/F/G/I protein . Its key specifications include:

The antibody has been validated for research applications including ELISA, Western Blot, and immunohistochemistry .

How should HIST1H2BC (Ab-15) Antibody be stored and handled?

For optimal performance and longevity of the HIST1H2BC antibody, researchers should follow these storage and handling guidelines:

• Store at -20°C for long-term storage

• For frequent use, aliquot to avoid repeated freeze-thaw cycles

• Maintain in a buffer containing preservatives (typically 0.03% Proclin 300)

• Store in 50% glycerol, 0.01M PBS, pH 7.4

• Avoid contamination

• Record date of reconstitution and number of freeze-thaw cycles

• Always centrifuge briefly before opening the vial after thawing

Proper storage and handling are critical for maintaining antibody activity and ensuring consistent experimental results.

What are the recommended applications and dilutions for HIST1H2BC (Ab-15) Antibody?

Based on available data, the HIST1H2BC (Ab-15) Antibody can be used in multiple applications with the following recommended dilutions:

These dilutions should be considered as starting points and may require optimization for your specific experimental conditions and sample types .

How can I validate the specificity of HIST1H2BC (Ab-15) Antibody in my experimental system?

Validating antibody specificity is crucial for ensuring reliable experimental results. For HIST1H2BC (Ab-15) Antibody, consider these validation approaches:

• Positive and negative controls: Use cell lines or tissues known to express or not express HIST1H2BC.

• Peptide competition assay: Pre-incubate the antibody with the immunizing peptide (peptide sequence around Lys-15) before application to your samples. This should abolish specific binding.

• Genetic knockdown validation: Compare antibody staining in wildtype cells versus cells with HIST1H2BC knockdown (siRNA) or knockout (CRISPR-Cas9).

• Multiple antibody comparison: Use another validated HIST1H2BC antibody (such as HIST1H2BC Ab-108 or Ab-120) that targets a different epitope and compare staining patterns .

• Mass spectrometry validation: For the ultimate confirmation, immunoprecipitate the protein and analyze by mass spectrometry to confirm identity.

Document all validation methods used as this strengthens the credibility of subsequent experimental results.

What is the relationship between HIST1H2BC and its related histone variants in chromatin research?

HIST1H2BC belongs to a family of histone H2B variants that share high sequence homology. When designing experiments, researchers should consider:

• Sequence conservation: HIST1H2BC shares significant sequence similarity with other H2B variants including H2BFH, H2BFA, H2BFK, and others (HIST1H2BE, HIST1H2BF, HIST1H2BG, HIST1H2BI) . This can affect antibody cross-reactivity.

• Functional redundancy: Due to high sequence similarity, these variants may have overlapping functions in chromatin structure and gene regulation.

• Tissue-specific expression: Different H2B variants may show tissue-specific or development-specific expression patterns.

• Distinguishing variants: When studying HIST1H2BC specifically, consider using techniques that can distinguish between highly similar variants:

  • Use antibodies against unique post-translational modifications

  • Employ RNA interference targeting unique 3'UTR sequences

  • Design PCR primers for variant-specific regions

How can I optimize the HIST1H2BC (Ab-15) Antibody for use in Chromatin Immunoprecipitation (ChIP) assays?

While not explicitly listed in the product information, polyclonal antibodies against histones are often suitable for ChIP applications with proper optimization:

• Crosslinking optimization: For histones, a shorter formaldehyde crosslinking time (5-10 minutes) may be sufficient due to their direct interaction with DNA.

• Chromatin fragmentation: Aim for fragments of 200-500bp for optimal resolution. Sonication parameters should be carefully optimized for your sample type.

• Antibody amount: Begin with 2-5μg of HIST1H2BC (Ab-15) Antibody per ChIP reaction and adjust based on results.

• Pre-clearing step: Include a pre-clearing step with protein A/G beads to reduce non-specific binding.

• Washing stringency: Optimize salt concentration in wash buffers to maintain specific binding while reducing background.

• Controls:

  • Input control (non-immunoprecipitated chromatin)

  • IgG control (normal rabbit IgG)

  • Positive control (antibody against a well-characterized histone mark)

• Validation: Confirm enrichment at known HIST1H2BC-associated genomic regions via qPCR before proceeding to sequencing.

What are the implications of HIST1H2BC in cancer research, particularly in breast cancer?

While the search results focus more on HIST1H2BD than HIST1H2BC specifically, the related findings provide valuable insights for HIST1H2BC research in cancer:

• Expression alterations: Similar to HIST1H2BD, HIST1H2BC expression may be dysregulated in cancer tissues. Studies have shown that HIST1H2BD is significantly upregulated in breast tumor cells compared to normal breast cells .

• Prognostic significance: Research has identified HIST1H2BD as a potential prognostic marker in breast cancer, with a hazard ratio of 0.712, indicating that higher expression is associated with improved survival . Similar studies may be warranted for HIST1H2BC.

• Subtype relevance: HIST1H2BD expression correlates with recurrence-free survival in specific breast cancer subtypes (luminal A, HER2+, and normal-like) . Researchers should investigate if HIST1H2BC shows similar subtype-specific patterns.

• Metastatic potential: Given that HIST1H2BD has been identified as a hub gene in metastatic breast cancer networks , HIST1H2BC may also play a role in metastatic processes.

• Therapeutic target potential: The identification of HIST1H2BD as a potential therapeutic target suggests that other histone variants including HIST1H2BC warrant investigation for similar potential.

Researchers studying HIST1H2BC in cancer contexts should consider these findings while designing experiments to elucidate its specific role.

How do post-translational modifications affect HIST1H2BC antibody recognition?

Histone proteins, including HIST1H2BC, undergo extensive post-translational modifications (PTMs) that can significantly impact antibody recognition:

• Epitope masking: The HIST1H2BC (Ab-15) Antibody targets the region around Lysine 15. This lysine can be modified by acetylation, methylation, or ubiquitination, potentially masking the epitope and preventing antibody binding.

• Modification-specific detection: When studying specific HIST1H2BC modifications, use antibodies that specifically recognize the modified form (e.g., anti-H2B-K15ac for acetylated K15).

• Sample preparation considerations:

  • For detecting total HIST1H2BC regardless of modifications, use denaturing conditions in Western blots

  • For studying specific modifications, avoid harsh detergents that might remove modifications

  • Consider using phosphatase or deacetylase inhibitors in lysis buffers to preserve modification states

• Cross-reactivity assessment: Validate that your HIST1H2BC (Ab-15) Antibody recognizes the unmodified form by comparing recognition patterns in samples treated with or without modifying enzymes.

• Modification profiling: Consider combining HIST1H2BC immunoprecipitation with mass spectrometry to profile all modifications present on the protein.

Understanding the modification landscape of your samples is essential for accurate interpretation of antibody-based detection results.

What are common troubleshooting strategies for weak or non-specific signals when using HIST1H2BC (Ab-15) Antibody?

When encountering issues with HIST1H2BC antibody performance, consider these methodological approaches:

• Weak or no signal:

  • Increase antibody concentration (decrease dilution)

  • Extend primary antibody incubation time (overnight at 4°C)

  • Enhance detection system (more sensitive substrate, signal amplification)

  • Check protein extraction protocol (histones require specific extraction methods)

  • Verify target protein expression in your sample

  • Ensure sample preparation preserves epitope accessibility

• Non-specific or high background:

  • Optimize blocking (try different blocking agents, increase blocking time)

  • Increase washing steps (more frequent, longer washes)

  • Decrease antibody concentration

  • Pre-adsorb antibody with similar proteins

  • Filter secondary antibody to remove aggregates

  • Use more stringent washing buffers

• Inconsistent results:

  • Standardize protein extraction and loading

  • Maintain consistent antibody lot numbers

  • Implement positive and loading controls

  • Document and standardize all protocol variables

• Protocol optimization for histone detection:

  • For Western blots, use specialized histone extraction buffers containing high salt

  • For IHC/IF, consider antigen retrieval methods optimized for nuclear proteins

  • Run SDS-PAGE with 15-18% gels for better resolution of small histone proteins

How can I design multiplexed immunoassays incorporating HIST1H2BC (Ab-15) Antibody?

Multiplexed assays allow simultaneous detection of multiple targets, providing valuable context for HIST1H2BC analysis:

• Antibody species selection:

  • Pair HIST1H2BC (Ab-15) rabbit polyclonal with antibodies raised in different species (mouse, goat, etc.)

  • Use species-specific secondary antibodies with distinct fluorophores

• Sequential immunostaining:

  • For antibodies from the same species, consider sequential detection with complete elution between rounds

  • Document order effects by alternating the sequence in control experiments

• Chromatic separation:

  • Use fluorophores with minimal spectral overlap

  • Include single-stained controls for spectral unmixing

• Complementary target selection:

  • Pair HIST1H2BC with other histones or chromatin-associated proteins

  • Include markers for specific cellular contexts (proliferation, differentiation, stress)

• Technical considerations:

  • Start with validated individual staining protocols before combining

  • Assess for antibody cross-reactivity in the multiplexed format

  • Consider tyramide signal amplification for weak signals

  • Optimize fixation to preserve all target epitopes

• Suggested combinations for chromatin research:

  • HIST1H2BC + histone modifications (H3K4me3, H3K27ac, etc.)

  • HIST1H2BC + chromatin remodeling factors

  • HIST1H2BC + transcription markers

What emerging technologies can enhance HIST1H2BC (Ab-15) Antibody applications in single-cell analysis?

Recent technological advances offer new opportunities for studying HIST1H2BC at the single-cell level:

• CyTOF/Mass cytometry:

  • Conjugate HIST1H2BC (Ab-15) Antibody with rare metal isotopes

  • Allows simultaneous detection of 40+ proteins with minimal spillover

  • Particularly valuable for heterogeneous tissue analysis

• Single-cell Western blot:

  • Apply microfluidic-based single-cell protein analysis

  • Can reveal cell-to-cell variation in HIST1H2BC expression

  • Useful for rare cell populations

• Imaging mass cytometry:

  • Combines immunohistochemistry with mass spectrometry

  • Provides spatial context for HIST1H2BC expression

  • High multiplexing capacity for complex microenvironment analysis

• Proximity ligation assay (PLA):

  • Detects protein-protein interactions involving HIST1H2BC

  • Provides information on spatial proximity (<40nm)

  • Can be combined with single-molecule FISH for protein-RNA associations

• CODEX (CO-Detection by indEXing):

  • Allows for highly multiplexed imaging (40+ proteins)

  • DNA-barcoded antibodies with iterative visualization

  • Preserves tissue architecture

These technologies can provide unprecedented insights into HIST1H2BC biology at single-cell resolution, enabling new discoveries in chromatin biology and epigenetic regulation.

How can HIST1H2BC (Ab-15) Antibody contribute to understanding epigenetic mechanisms in development and disease?

HIST1H2BC (Ab-15) Antibody enables investigation of several key aspects of epigenetic regulation:

• Chromatin structure dynamics:

  • Track HIST1H2BC incorporation during cell cycle progression

  • Investigate its role in chromatin compaction/decompaction

  • Study its distribution in euchromatin versus heterochromatin regions

• Histone exchange mechanisms:

  • Examine HIST1H2BC turnover rates in different genomic contexts

  • Compare with replication-independent histone variants

  • Investigate factors regulating HIST1H2BC deposition and removal

• Disease-specific alterations:

  • Research has identified histone variants as potential diagnostic and prognostic markers in cancer

  • HIST1H2BD (related variant) shows prognostic value in breast cancer with a hazard ratio of 0.712

  • Investigation of HIST1H2BC alterations across cancer types may reveal similar patterns

• Therapeutic targeting:

  • Study how epigenetic drugs affect HIST1H2BC incorporation and modifications

  • Explore correlations between treatment response and HIST1H2BC status

  • Assess potential of HIST1H2BC as a biomarker for epigenetic therapy response

• Developmental programming:

  • Track changes in HIST1H2BC dynamics during cellular differentiation

  • Investigate its role in establishing and maintaining cell type-specific gene expression

  • Study inheritance patterns of HIST1H2BC-associated chromatin states

The antibody provides a valuable tool for interrogating these complex epigenetic mechanisms across diverse biological contexts.