SPCC1235.17 Antibody

What is SPCC1235.17 and what is its functional role in Schizosaccharomyces pombe?

SPCC1235.17 is a protein expressed in Schizosaccharomyces pombe (fission yeast), identified by UniProt Number G2TRT7. The protein is typically studied in the strain 972 / ATCC 24843 context, which serves as the standard laboratory reference strain for fission yeast research. While specific functional characterization research continues, antibodies against this protein enable researchers to investigate its expression patterns, cellular localization, and potential interaction partners through various immunological techniques. The protein's structural and functional analysis is facilitated by specific antibody detection in both native and experimental conditions .

What applications are most suitable for SPCC1235.17 antibody?

SPCC1235.17 antibody has been validated for two primary applications:

*Optimal dilutions should be determined empirically for each experimental system

The polyclonal nature of commercially available SPCC1235.17 antibodies makes them particularly suitable for these applications, as they recognize multiple epitopes on the target protein, potentially increasing detection sensitivity. When designing experiments, researchers should consider that this antibody is typically supplied unconjugated and requires appropriate secondary detection systems .

What is the recommended storage protocol for maintaining SPCC1235.17 antibody activity?

For optimal preservation of antibody activity, SPCC1235.17 antibody should be stored at either -20°C or -80°C according to manufacturer specifications. Proper storage considerations include:

• Aliquoting the antibody upon receipt to minimize freeze-thaw cycles

• Storing working dilutions at 4°C for short-term use (1-2 weeks)

• Avoiding repeated freeze-thaw cycles that can lead to antibody denaturation and reduced binding efficacy

• Maintaining sterile conditions to prevent microbial contamination

Storage buffer composition may include glycerol as a cryoprotectant, though specific formulations may vary between suppliers. When preparing working dilutions, researchers should use buffers that maintain antibody stability and activity while reducing non-specific binding .

How can I validate the specificity of SPCC1235.17 antibody for my research?

Validating antibody specificity is essential for generating reliable research data. For SPCC1235.17 antibody, consider implementing these validation approaches:

• Positive control validation: Use the supplied antigen (200μg) as a positive control in your detection system to confirm antibody binding to the target protein .

• Negative control testing: Employ the provided pre-immune serum (1ml) as a negative control to assess background and non-specific binding .

• Knockout/knockdown verification: If available, test the antibody against SPCC1235.17-deficient yeast strains to confirm signal specificity.

• Western blot molecular weight verification: Confirm detection of a band at the predicted molecular weight for SPCC1235.17.

• Peptide competition assay: Pre-incubate the antibody with excess target peptide to demonstrate signal elimination in subsequent detection.

This multi-faceted approach ensures confidence in experimental results and helps identify potential cross-reactivity issues before proceeding with complex experiments.

How can I optimize SPCC1235.17 antibody performance in non-standard experimental conditions?

Optimizing SPCC1235.17 antibody performance in challenging experimental conditions requires methodical adjustment of several parameters:

• Buffer optimization: For detecting native conformations, test phosphate, Tris, and HEPES buffer systems at various pH ranges (6.5-8.0) to identify conditions that maximize signal-to-noise ratio.

• Detergent selection: When working with membrane fractions, evaluate different detergent concentrations (common options: 0.1-0.5% Triton X-100, 0.05-0.2% Tween-20, or 0.1-1% NP-40) to improve protein extraction while preserving epitope accessibility.

• Blocking agent testing: Different blocking agents (BSA, non-fat milk, casein, or commercial alternatives) can significantly impact antibody performance in specific applications.

• Cross-linking considerations: If performing cross-linking immunoprecipitation, optimize formaldehyde or other cross-linker concentrations (typically 0.1-1%) and exposure times to preserve protein-protein interactions without masking epitopes.

• Temperature modifications: While standard incubations are typically performed at room temperature or 4°C, testing a range of temperatures may improve binding kinetics for certain applications.

Each optimization step should include appropriate controls and be performed systematically, changing one variable at a time to identify optimal conditions for your specific experimental system .

What computational approaches can improve SPCC1235.17 antibody design and application?

Advanced computational methods, particularly those developed for antibody design, can significantly enhance SPCC1235.17 antibody performance. The RosettaAntibodyDesign (RAbD) framework offers powerful approaches for researchers seeking to optimize their antibody-based experiments:

• Epitope analysis and optimization: Computational prediction of optimal epitopes on SPCC1235.17 can guide the development of more specific antibodies. RAbD samples the diverse sequence, structure, and binding space of antibody-antigen complexes to identify potential improvements .

• Affinity maturation in silico: Before undertaking expensive and time-consuming experimental affinity maturation, researchers can use computational methods to predict mutations likely to enhance binding affinity. This approach works by sampling CDR (Complementarity-Determining Region) structures from databases of canonical clusters .

• Cross-reactivity prediction: Computational methods can identify potential cross-reactive epitopes in other yeast proteins, helping researchers anticipate and mitigate specificity issues.

• Structure-guided experimental design: When crystal structures are unavailable, antibody modeling can guide experimental design by predicting epitope accessibility in different experimental conditions.

• CDR optimization: RAbD can be used to redesign single or multiple CDRs with loops of different length, conformation, and sequence to potentially improve binding characteristics .

Implementing these computational approaches requires interdisciplinary collaboration but can significantly reduce experimental iterations and improve antibody performance in challenging applications.

What strategies should I employ when troubleshooting inconsistent SPCC1235.17 antibody results?

When encountering inconsistent results with SPCC1235.17 antibody, implement a systematic troubleshooting approach:

• Antibody quality assessment:

  • Verify antibody integrity by SDS-PAGE (look for expected IgG bands at ~50kDa and ~25kDa)

  • Test a new aliquot to rule out degradation from improper storage or handling

  • Confirm antibody concentration using absorbance at 280nm

• Protocol examination:

  • Review each step of your experimental protocol for deviations

  • Evaluate reagent quality and preparation dates

  • Consider temperature fluctuations during critical incubation steps

• Sample preparation analysis:

  • Verify protein extraction efficiency from yeast cells

  • Check for protease inhibitor effectiveness

  • Assess protein concentration methods for consistency

• Methodological modifications:

  • Adjust antibody concentration through systematic titration experiments

  • Modify incubation times and temperatures

  • Test different detection systems (fluorescent vs. chemiluminescent for Western blots)

• Instrument calibration:

  • Ensure proper calibration of imaging systems

  • Verify consistent exposure parameters

  • Compare results across different detection platforms if available

Document all troubleshooting steps methodically to identify patterns that may reveal the source of inconsistency. Consider using the supplied positive control (200μg antigen) alongside experimental samples to provide a consistent reference point across experiments .

How does SPCC1235.17 protein expression vary across Schizosaccharomyces pombe growth phases?

Expression analysis of SPCC1235.17 across different growth phases requires careful experimental design. While specific expression data for this protein varies across research laboratories, a methodological approach to characterizing its expression includes:

• Time-course sampling protocol:

  • Inoculate S. pombe cultures and measure OD600 to track growth phases

  • Collect samples at consistent intervals spanning lag, exponential, and stationary phases

  • Process samples immediately or flash-freeze to preserve protein integrity

• Quantitative Western blot analysis:

  • Use consistent protein extraction methods across all time points

  • Load equal total protein amounts (typically 20-50μg) per lane

  • Include housekeeping protein controls (e.g., tubulin or actin) for normalization

  • Employ the SPCC1235.17 antibody at optimized dilutions (1:500-1:2000)

• Expression data analysis:

  • Quantify band intensities using image analysis software

  • Normalize to housekeeping protein expression

  • Plot relative expression levels across time points

  • Perform statistical analysis to identify significant changes

• Validation approaches:

  • Complement protein expression data with RT-qPCR for mRNA levels

  • Consider fluorescent reporter constructs for live-cell imaging if appropriate

This methodological framework provides robust data on SPCC1235.17 expression patterns that can inform experimental design and interpretation of functional studies .

How can I apply SPCC1235.17 antibody in complex experimental systems beyond standard applications?

Extending SPCC1235.17 antibody applications beyond standard ELISA and Western blot techniques requires creative adaptation of immunological methods:

• Chromatin Immunoprecipitation (ChIP):

  • If SPCC1235.17 has potential DNA-binding activity, optimize cross-linking conditions (typically 1% formaldehyde for 10-15 minutes)

  • Sonicate chromatin to 200-500bp fragments

  • Use 2-5μg antibody per ChIP reaction

  • Validate enrichment with qPCR of predicted binding regions

• Proximity Ligation Assay (PLA):

  • For detecting protein-protein interactions in situ

  • Combine SPCC1235.17 antibody with antibodies against suspected interaction partners

  • Requires species-specific secondary antibodies and optimization of fixation protocols

• Super-resolution microscopy:

  • Adapt immunofluorescence protocols for STORM or PALM imaging

  • Test different fixation methods to preserve epitope accessibility

  • Consider direct fluorophore conjugation to minimize spatial displacement

• Mass spectrometry-based interaction studies:

  • Optimize immunoprecipitation conditions for downstream mass spectrometry

  • Consider chemical cross-linking to stabilize transient interactions

  • Implement appropriate controls to distinguish specific from non-specific interactors

• Single-cell analysis applications:

  • Adapt protocols for flow cytometry or mass cytometry

  • Optimize permeabilization conditions for intracellular detection

  • Consider fluorescent secondary antibody selection for multiplexing capabilities

For each advanced application, preliminary optimization experiments are essential, leveraging the provided positive control antigen to establish baseline parameters before proceeding to complex biological samples .