RBKS Antibody

What is RBKS Antibody and what protein does it target?

RBKS antibody targets the human ribokinase protein (also known as RBKS, EC 2.7.1.15), which is encoded by the RBKS gene . This antibody is commonly available as a polyclonal rabbit IgG that has been antigen and protein A affinity-purified . The specific antibody referenced in the search results is raised against purified recombinant Human RBKS protein (Accession#: NP_071411.1; Met1-Phe322) . Ribokinase is an enzyme involved in carbohydrate metabolism, specifically in the phosphorylation of ribose to ribose-5-phosphate in the pentose phosphate pathway.

What are the common applications for RBKS Antibody?

RBKS antibody can be utilized across multiple experimental platforms, with the most common applications including:

• Western Blot: For detecting RBKS protein in cell or tissue lysates

• Immunohistochemistry-Paraffin (IHC-P): For visualizing RBKS localization in fixed tissue sections

• Immunoprecipitation (IP): For isolating RBKS protein and its complexes

• Enzyme-Linked Immunosorbent Assay (ELISA): For quantitative detection of RBKS protein

The optimal dilution for each application must be experimentally determined based on the specific experimental conditions and the particular antibody being used .

What validation controls should be used when working with RBKS Antibody?

Proper validation controls are essential for ensuring reliable results with RBKS antibody. Recommended controls include:

• Positive control: Cell lines or tissues known to express RBKS

• Negative control: RBKS knockout (KO) cell lines or tissues, which represent the gold standard for specificity testing

• Isotype control: Use of non-specific antibodies of the same isotype (e.g., rabbit IgG) to identify non-specific binding

• Blocking peptide: Competition assays using the immunizing peptide to confirm specificity

• Secondary antibody-only control: To identify background from the detection system

How should RBKS Antibody be stored and handled for optimal performance?

For optimal performance and longevity of the RBKS antibody:

• Store at 4°C in the dark, as recommended for the biotin-conjugated version

• Avoid repeated freeze-thaw cycles

• Use sterile techniques when handling

• Follow manufacturer's recommendations for reconstitution if lyophilized

• Consider aliquoting the antibody to minimize freeze-thaw cycles if stored at -20°C

• Verify stability period (typically guaranteed for 1 year from date of receipt)

What information should I record when using RBKS Antibody?

To ensure reproducibility, detailed documentation is crucial when using RBKS antibody. Record:

Scientific papers must include detailed methods sections, as well as unambiguous descriptions of the antibodies used including RRID, source, catalog number, details as to the type of antibody, immunogen used to raise the antibody, and protein concentrations used in each assay .

How can I validate RBKS Antibody specificity for my specific application?

Antibody performance is strongly influenced by assay context, requiring validation specific to each application . For RBKS antibody:

Western Blot Validation:

• Test on cell lines with known RBKS expression levels

• Include RBKS knockout or knockdown samples as negative controls

• Verify the observed molecular weight matches the predicted size of RBKS

• Perform peptide competition assay with the immunizing peptide

• Compare results with another RBKS antibody targeting a different epitope

IHC-P Validation:

• Test on tissues with known RBKS expression patterns

• Compare with in situ hybridization data for RBKS mRNA

• Include RBKS knockout tissues as negative controls

• Evaluate specificity through peptide competition

• Verify subcellular localization is consistent with known RBKS biology

Because the performance of primary antibodies is strongly influenced by assay context, validation strategies must be tailored to each type of immunoassay .

How can I troubleshoot inconsistent results when using RBKS Antibody in Western blotting?

Inconsistent Western blot results with RBKS antibody may stem from multiple sources:

• Sample Preparation Issues:

  • Ensure complete protein denaturation and reduction

  • Verify protein extraction method is appropriate for RBKS

  • Check for protease inhibitor inclusion in lysis buffers

  • Quantify and normalize protein loading accurately

• Antibody-Related Factors:

  • Optimize primary antibody concentration (perform titration)

  • Ensure antibody lot consistency (lot-to-lot variation can be significant)

  • Verify storage conditions haven't compromised antibody activity

  • Consider testing alternative RBKS antibodies targeting different epitopes

• Protocol Optimization:

  • Adjust blocking conditions to reduce background

  • Optimize incubation times and temperatures

  • Modify washing steps to improve signal-to-noise ratio

  • Consider transfer efficiency for RBKS protein molecular weight

• Detection System:

  • Ensure appropriate secondary antibody selection

  • Optimize ECL substrate or fluorescent detection parameters

  • Check for scanner/imager sensitivity settings

The validation of primary antibodies is crucial as their performance is strongly influenced by assay context .

What are the considerations for cross-reactivity assessment of RBKS Antibody?

Assessing potential cross-reactivity of RBKS antibody is critical for accurate data interpretation:

• Sequence Homology Analysis:

  • Identify proteins with sequence similarity to RBKS

  • Assess whether the immunogen sequence has homology to other proteins

• Experimental Assessment:

  • Test on samples expressing related proteins but not RBKS

  • Use mass spectrometry to identify all proteins immunoprecipitated by the antibody

  • Perform epitope mapping to understand antibody binding sites

• Species Cross-Reactivity:

  • If using across species, align RBKS sequences to predict cross-reactivity

  • Experimentally verify cross-reactivity with each species

  • Note that the example RBKS antibody is specified for human reactivity

• Documentation:

  • Record all cross-reactivity testing in lab notebooks

  • Report observed cross-reactivity in publications

  • Contact manufacturers to report unexpected cross-reactivity

How does the performance of polyclonal vs. monoclonal RBKS Antibodies compare?

The choice between polyclonal and monoclonal RBKS antibodies depends on research goals:

Polyclonal RBKS Antibodies:

• Recognize multiple epitopes, potentially increasing detection sensitivity

• May have greater batch-to-batch variability

• Can be more robust against minor protein modifications or conformational changes

• Example: The referenced biotin-conjugated polyclonal rabbit IgG RBKS antibody

Monoclonal RBKS Antibodies:

• Recognize a single epitope, providing higher specificity

• Offer greater consistency between batches

• May be more sensitive to epitope masking or modification

• Preferred for applications requiring high specificity

Researchers are encouraged, where possible, to avoid polyclonals and use well-characterized recombinant antibodies instead for improved reproducibility .

What approaches can help resolve contradictory results with RBKS Antibody across different studies?

When faced with contradictory results using RBKS antibody:

• Antibody Identity Verification:

  • Confirm RBKS antibodies are truly different by checking RRIDs or sequences

  • Some antibodies may have different RRIDs but actually be the same clone under different names

  • Verify lot numbers, especially for polyclonal antibodies

• Methodological Differences:

  • Compare detailed protocols including sample preparation, dilutions, and detection methods

  • Standardize key experimental parameters across studies

  • Implement the same positive and negative controls

• Orthogonal Validation:

  • Confirm findings using alternative methods not dependent on antibodies

  • Employ genetic approaches (knockout/knockdown) to verify specificity

  • Consider using mass spectrometry for protein identification

• Multi-laboratory Validation:

  • Establish collaborations to test the same RBKS antibody under different conditions

  • Document and share detailed protocols to identify sources of variation

How should I design Western blot experiments using RBKS Antibody?

Optimal Western blot design for RBKS antibody requires careful consideration of multiple factors:

• Sample Preparation:

  • Select appropriate lysis buffer (considering RBKS subcellular localization)

  • Include protease and phosphatase inhibitors to prevent degradation

  • Determine optimal protein concentration (typically 20-50 μg per lane)

  • Properly denature samples (95°C for 5 minutes in reducing buffer)

• Gel Selection and Separation:

  • Choose appropriate percentage gel based on RBKS molecular weight

  • Consider gradient gels for better resolution

  • Ensure complete protein transfer to membrane (optimize transfer time/voltage)

• Antibody Application:

  • Determine optimal dilution experimentally for the RBKS antibody

  • Use appropriate blocking buffer to minimize background

  • Optimize incubation time and temperature

  • Include positive control (lysate known to express RBKS)

  • Include negative control (RBKS knockout or knockdown)

• Detection and Quantification:

  • Select appropriate detection system compatible with biotin conjugation

  • Ensure linear range of detection for quantification

  • Use appropriate normalization controls (loading controls)

  • Document exposure settings for reproducibility

Practical strategies for the validation of primary antibodies are essential since their performance is strongly influenced by assay context .

What are the critical parameters for immunohistochemistry with RBKS Antibody?

For successful IHC with RBKS antibody, consider:

• Tissue Preparation:

  • Optimize fixation method and duration

  • Select appropriate embedding and sectioning techniques

  • Determine need for antigen retrieval methods

  • Establish optimal section thickness

• Antibody Application:

  • Titrate antibody concentration to determine optimal dilution

  • Select appropriate diluent to minimize background

  • Optimize incubation time, temperature, and washing conditions

  • Include appropriate blocking steps

• Detection System:

  • Choose detection system compatible with biotin-conjugated antibody

  • Consider amplification methods if RBKS expression is low

  • Select appropriate counterstains for cellular context

  • Establish image acquisition parameters

• Controls:

  • Include positive control tissue with known RBKS expression

  • Include negative control tissue (RBKS knockout when available)

  • Use isotype control antibody to assess non-specific binding

  • Include secondary-only control to assess background

How can I optimize immunoprecipitation protocols using RBKS Antibody?

Effective immunoprecipitation with RBKS antibody requires:

• Lysis Conditions:

  • Select lysis buffer that preserves protein-protein interactions

  • Include appropriate protease and phosphatase inhibitors

  • Optimize cell lysis conditions (temperature, duration, method)

  • Clear lysates thoroughly before immunoprecipitation

• Antibody Binding:

  • Determine optimal amount of RBKS antibody

  • Select appropriate beads considering the biotin conjugation

  • Optimize antibody-bead binding conditions

  • Determine optimal incubation time with sample

• Washing and Elution:

  • Establish washing stringency to minimize non-specific binding

  • Optimize washing buffer composition and number of washes

  • Select appropriate elution conditions

  • Verify successful elution of target protein

• Controls and Validation:

  • Include IgG control immunoprecipitation

  • Verify specific enrichment of RBKS in immunoprecipitates

  • Consider confirming interacting partners by mass spectrometry

  • Validate key interactions by reciprocal immunoprecipitation

What considerations are important when using RBKS Antibody for ELISA?

For optimal ELISA performance with RBKS antibody:

• Assay Format Selection:

  • Determine appropriate ELISA format (direct, indirect, sandwich, competitive)

  • For sandwich ELISA, pair RBKS antibody with a complementary antibody

  • Consider whether biotin conjugation can be leveraged in the detection system

• Antibody and Reagent Optimization:

  • Determine optimal antibody concentration through titration

  • Optimize blocking buffer to minimize background

  • Select appropriate detection system

  • Establish standard curve parameters if quantitative analysis is needed

• Sample Preparation:

  • Determine optimal sample dilution to remain in the linear range

  • Consider sample pre-treatments to expose epitopes

  • Establish appropriate controls for matrix effects

• Validation and Controls:

  • Include standard curves with recombinant RBKS protein

  • Establish assay specificity using RBKS-depleted samples

  • Determine assay sensitivity, precision, and reproducibility

  • Include appropriate negative and positive controls

How can I quantify Western blot results using RBKS Antibody?

Accurate quantification of RBKS protein by Western blot requires:

• Image Acquisition:

  • Capture images within the linear dynamic range of the detection system

  • Use consistent exposure settings between experimental replicates

  • Avoid pixel saturation which compromises quantification

  • Include a standard curve if absolute quantification is required

• Normalization Approaches:

  • Use appropriate loading controls (housekeeping proteins)

  • Consider total protein normalization methods (e.g., stain-free technology)

  • Validate stability of reference proteins under experimental conditions

  • Apply consistent normalization across all samples

• Quantification Methods:

  • Use appropriate software for densitometry analysis

  • Define bands consistently across all samples

  • Subtract background appropriately

  • Apply statistical tests appropriate for the experimental design

• Data Representation:

  • Present raw blot images alongside quantification

  • Report both technical and biological replicates

  • Include error bars and statistical significance

  • Present fold-changes relative to controls when appropriate

What are common sources of variability in RBKS Antibody experiments?

Understanding and controlling sources of variability is crucial:

• Antibody-Related Variability:

  • Lot-to-lot variations, especially for polyclonal antibodies

  • Antibody degradation due to improper storage

  • Inconsistent dilution preparation

  • Cross-reactivity with similar proteins

• Sample-Related Variability:

  • Inconsistent sample preparation

  • Protein degradation during storage

  • Variation in protein extraction efficiency

  • Heterogeneity in biological samples

• Protocol-Related Variability:

  • Inconsistent blocking or washing steps

  • Temperature and incubation time variations

  • Differences in detection reagents or equipment settings

  • Variations in quantification methods

• Researcher-Related Variability:

  • Differences in technique between researchers

  • Inconsistent application of protocols

  • Unconscious bias in data analysis

  • Variations in reporting practices

It has been estimated that ~50% of commercial antibodies fail to meet even basic standards for characterization, resulting in financial losses of $0.4–1.8 billion per year in the United States alone .

How should I report RBKS Antibody usage in publications to enhance reproducibility?

Comprehensive reporting enhances experimental reproducibility:

• Antibody Identification:

  • Include RRID (Research Resource Identifier)

  • Provide manufacturer, catalog number, and lot number

  • Specify antibody type (polyclonal/monoclonal) and host species

  • Note conjugation (e.g., biotin)

• Validation Evidence:

  • Describe validation methods used

  • Include or reference validation data

  • Specify positive and negative controls

  • Report known limitations or cross-reactivity

• Experimental Conditions:

  • Detail buffer compositions

  • Specify dilutions used for each application

  • Report incubation conditions (time, temperature)

  • Describe blocking and washing procedures

• Data Analysis Methods:

  • Explain quantification approach

  • Describe normalization method

  • Specify software used for analysis

  • Detail statistical methods applied

Scientific papers must include detailed methods sections, as well as unambiguous descriptions of the antibodies used (RRID, source, catalog number, details as to the type of antibody, immunogen used to raise the antibody, protein concentrations used in each assay) .

What advantages do recombinant RBKS Antibodies offer over traditional antibodies?

Recombinant antibody technology provides several advantages:

• Consistency and Reproducibility:

  • Defined amino acid sequence eliminates lot-to-lot variation

  • Consistent performance across experiments and time

  • Reduced risk of antibody drift over production cycles

• Customization Potential:

  • Ability to engineer specific characteristics

  • Optimization of affinity and specificity

  • Creation of novel formats (e.g., bispecific antibodies)

• Ethical and Practical Benefits:

  • Reduction in animal use for antibody production

  • Potential for unlimited supply

  • Known molecular identity allowing better cross-validation

• Research Transparency:

  • Sequenced antibodies enable true validation between laboratories

  • Prevention of mistaken "cross-validation" with differently named but identical antibodies

  • Enhanced reproducibility through exact knowledge of molecular tools

Where possible, end users should avoid the use of polyclonals, and use well-characterized recombinant antibodies instead for improved reproducibility .

How is RBKS Antibody research contributing to our understanding of metabolic pathways?

RBKS antibody enables investigation of ribokinase's role in metabolism:

• Pentose Phosphate Pathway Regulation:

  • Visualization of RBKS expression patterns across tissues

  • Correlation of RBKS levels with metabolic states

  • Analysis of RBKS interactions with regulatory proteins

• Disease Associations:

  • Examination of RBKS expression in metabolic disorders

  • Investigation of potential dysregulation in cancer metabolism

  • Study of RBKS in rare genetic disorders affecting ribose metabolism

• Cellular Localization Studies:

  • Determination of subcellular compartmentalization of RBKS

  • Analysis of dynamic changes in localization under metabolic stress

  • Co-localization with other metabolic enzymes

• Post-translational Modifications:

  • Detection of regulatory modifications of RBKS protein

  • Correlation of modifications with enzymatic activity

  • Identification of signaling pathways controlling RBKS function

What role can RBKS Antibody play in advancing multi-omic research approaches?

Integration of RBKS antibody-based methods with other technologies:

• Proteomics Integration:

  • RBKS immunoprecipitation coupled with mass spectrometry

  • Correlation of RBKS protein levels with global proteome changes

  • Analysis of RBKS interaction networks

• Metabolomics Connections:

  • Relating RBKS expression to metabolite profiles

  • Understanding impact of RBKS activity on metabolic fluxes

  • Integrating enzyme levels with pathway outputs

• Transcriptomics Correlation:

  • Comparing RBKS protein levels with mRNA expression

  • Investigating post-transcriptional regulation

  • Analyzing coordinated expression with related enzymes

• Single-cell Multi-omics:

  • Using RBKS antibody in single-cell protein analysis

  • Correlating RBKS expression with single-cell transcriptomics

  • Investigating cell-to-cell variability in metabolic enzyme expression