SPAC1F7.10 Antibody

What is SPAC1F7.10 and why is it significant in fission yeast research?

SPAC1F7.10 refers to a specific gene locus in Schizosaccharomyces pombe (fission yeast) genome. This gene is part of the systematic naming convention used for S. pombe, where SPAC indicates the chromosome location, followed by specific identifiers . The significance of studying this gene lies in understanding fission yeast biology, particularly as S. pombe serves as an important model organism for eukaryotic cell biology research. While the specific function of SPAC1F7.10 isn't explicitly detailed in the provided search results, it should be noted that genes in similar chromosomal regions, such as SPAC1F7.05 (cdc22), are known to be cell cycle-regulated genes involved in G1-S phase transitions .

Understanding SPAC1F7.10 contributes to our broader knowledge of gene regulation patterns in fission yeast, which has significant implications for understanding conserved biological processes across eukaryotes. Research into fission yeast genes has historically provided valuable insights into fundamental cellular processes, making antibodies against these gene products critical research tools .

What basic applications is the SPAC1F7.10 Antibody suitable for?

The SPAC1F7.10 Antibody is validated for two primary applications in basic research:

• Western Blotting (WB): The antibody can be used to detect and quantify the SPAC1F7.10 protein in cell lysates or protein extracts from S. pombe. This enables researchers to study protein expression levels under various experimental conditions .

• Enzyme-Linked Immunosorbent Assay (ELISA): The antibody can be utilized in ELISA protocols to detect and quantify the target protein in solution-based samples .

Both applications are fundamental techniques in molecular biology research that allow for the identification and relative quantification of the target protein. The antibody has been specifically tested to ensure proper identification of the antigen in these applications . For researchers new to working with fission yeast proteins, these applications provide essential starting points for characterizing SPAC1F7.10 protein expression and regulation.

What is the optimal storage condition for SPAC1F7.10 Antibody?

The SPAC1F7.10 Antibody should be stored at -20°C or -80°C upon receipt to maintain its functionality and specificity. Researchers should avoid repeated freeze-thaw cycles as these can degrade antibody quality and reduce binding efficiency . The antibody is supplied in liquid form with a storage buffer containing 0.03% Proclin 300 as a preservative, 50% Glycerol, and 0.01M PBS at pH 7.4 .

The high glycerol content (50%) helps prevent freezing damage during storage at -20°C, which is important for maintaining the antibody's tertiary structure and binding capacity. For long-term storage, -80°C is recommended, while -20°C is suitable for antibodies that will be used within a few months. When handling the antibody, it should be kept on ice during experiments and returned to appropriate storage conditions promptly after use to maximize shelf life and performance consistency .

How should researchers design experiments to validate SPAC1F7.10 Antibody specificity in S. pombe?

Validating antibody specificity is crucial for ensuring reliable results in fission yeast research. Researchers should implement a multi-step validation strategy:

• Positive and Negative Controls: Include wild-type S. pombe extracts as positive controls and either knockout strains (ΔSPAC1F7.10) or heterologous species as negative controls to confirm specificity .

• Peptide Competition Assay: Pre-incubate the antibody with purified recombinant SPAC1F7.10 protein (the immunogen) before applying to samples. Reduction or elimination of signal confirms specificity .

• Molecular Weight Verification: Ensure the detected protein band in Western blots matches the predicted molecular weight of SPAC1F7.10 protein.

• Cross-reactivity Assessment: Test the antibody against protein extracts from related yeast species to evaluate potential cross-reactivity, keeping in mind that this antibody is specifically raised against S. pombe strain 972/ATCC 24843 .

• Technical Replicates: Perform multiple independent experiments to confirm consistent performance across different sample preparations and experimental conditions.

For cell cycle studies, researchers should consider synchronizing cells using techniques like elutriation or temperature-sensitive cell cycle mutants, similar to methods used in other fission yeast studies . This approach allows for observation of protein expression patterns throughout different cell cycle phases and can provide valuable context for interpreting antibody specificity results.

How does SPAC1F7.10 Antibody compare to other antibodies for fission yeast research?

When comparing SPAC1F7.10 Antibody to other antibodies used in fission yeast research, several factors should be considered:

• Antibody Type: The SPAC1F7.10 Antibody is a polyclonal antibody, which typically provides higher sensitivity but potentially lower specificity compared to monoclonal alternatives . This is advantageous for detecting proteins expressed at low levels but may require more extensive validation to confirm specificity.

• Species Reactivity: This antibody is specifically developed for Schizosaccharomyces pombe (strain 972/ATCC 24843), making it highly specialized for fission yeast research . Unlike broader-spectrum antibodies that may cross-react with multiple species, this focused reactivity provides greater confidence in detecting the intended target in fission yeast studies.

• Production Method: The antibody is antigen-affinity purified, which enhances its specificity compared to crude serum antibodies often used in research . The purification process removes potentially cross-reactive antibodies, resulting in lower background and cleaner results.

• Applications: The antibody is validated for ELISA and Western blotting, which covers the most common applications in protein research . This range is typical for fission yeast antibodies, though some antibodies for well-studied proteins may offer additional validated applications like immunofluorescence or immunoprecipitation.

The made-to-order nature (14-16 weeks lead time) suggests this is a specialized research tool rather than a commonly stocked antibody, reflecting the relatively specialized nature of SPAC1F7.10 research compared to more widely studied fission yeast proteins like those involved in cell cycle regulation (e.g., cdc22) .

How can SPAC1F7.10 Antibody be used to study cell cycle regulation in fission yeast?

To utilize SPAC1F7.10 Antibody in cell cycle regulation studies, researchers should implement synchronized cell culture techniques similar to those employed in comprehensive cell cycle gene expression studies:

• Synchronization Methods: Employ either elutriation (physical separation based on cell size) or temperature-sensitive cell cycle mutants (such as cdc10-V50) to obtain synchronized cell populations . This allows for precise sampling at specific cell cycle stages.

• Time-Course Western Blotting: After synchronization, collect samples at regular intervals (e.g., 15-minute intervals for 4-5 hours) and perform Western blotting with SPAC1F7.10 Antibody to track protein expression changes throughout the cell cycle .

• Correlation Analysis: Compare SPAC1F7.10 protein expression patterns with known cell cycle markers, such as those listed in the comprehensive cell cycle gene expression dataset (e.g., cdc22 for G1-S, cdc25 for G2) . This table provides valuable reference points:

• Promoter Analysis: If SPAC1F7.10 shows cell cycle-regulated expression, analyze its promoter region for common cell cycle-specific motifs such as MCB, SFF, or ACE2 . This can provide insights into its transcriptional regulation mechanisms.

• Genetic Perturbation: Combine antibody-based protein detection with genetic manipulations (e.g., deletion mutants, overexpression strains) to analyze functional relationships between SPAC1F7.10 and known cell cycle regulators .

The comprehensive approach used for identifying 747 cell cycle-regulated genes in fission yeast provides an excellent methodological framework that can be adapted for studying potential cell cycle roles of SPAC1F7.10 .

What methodologies allow for quantitative analysis of SPAC1F7.10 protein levels using this antibody?

For quantitative analysis of SPAC1F7.10 protein levels, researchers should employ several complementary methodologies:

• Quantitative Western Blotting: Utilize chemiluminescent or fluorescent detection systems with the SPAC1F7.10 Antibody, coupled with appropriate loading controls (e.g., tubulin or actin) . Image analysis software can then be used to quantify relative band intensities across samples. Important considerations include:

  • Establishing a linear detection range through serial dilutions of samples

  • Consistent antibody dilutions and incubation times

  • Multiple technical and biological replicates

• Quantitative ELISA: Develop a sandwich or direct ELISA protocol using the SPAC1F7.10 Antibody to enable absolute quantification of protein levels . This requires:

  • Generation of a standard curve using purified recombinant SPAC1F7.10 protein

  • Optimization of antibody concentrations and blocking conditions

  • Statistical analysis of replicate measurements

• Data Normalization: When analyzing time-course experiments (such as cell cycle studies), implement robust normalization strategies similar to those used in gene expression studies:

  • Missing value replacement using neighbors' average values

  • Application of appropriate statistical transformations for proper clustering

• False Discovery Rate Estimation: For complex experimental designs with multiple comparisons, estimate the false discovery rate using randomized data approaches as described for gene expression studies . This enhances the statistical reliability of results.

• Integration with Transcriptomic Data: For comprehensive understanding, consider correlating protein-level measurements with existing transcriptomic datasets, using data processing methods similar to those employed for microarray analysis in cell cycle studies .

These methodologies provide a robust framework for generating reliable quantitative data regarding SPAC1F7.10 protein expression under various experimental conditions.

How can researchers investigate potential interactions between SPAC1F7.10 and other fission yeast proteins?

Investigating protein-protein interactions involving SPAC1F7.10 requires sophisticated methodological approaches beyond basic antibody applications:

• Co-Immunoprecipitation (Co-IP): While the SPAC1F7.10 Antibody is not explicitly validated for immunoprecipitation, researchers might test its capability for pulling down protein complexes:

  • Crosslink the antibody to appropriate beads (Protein A/G or magnetic)

  • Optimize lysis conditions to preserve native protein interactions

  • Identify co-precipitated proteins through mass spectrometry

  • Validate interactions with reciprocal Co-IPs using antibodies against identified partners

• Proximity-Based Labeling: Employ BioID or APEX2 fusion strategies with SPAC1F7.10 to identify proximal proteins, then verify these interactions using the SPAC1F7.10 Antibody in confirmatory Western blot experiments.

• Yeast Two-Hybrid Screening: Perform Y2H screens using SPAC1F7.10 as bait, followed by validation of identified interactions through other methods including the SPAC1F7.10 Antibody .

• Co-localization Studies: If the antibody can be optimized for immunofluorescence, assess co-localization with known cell cycle regulators or other proteins of interest throughout the cell cycle.

• Comparative Analysis with Known Interaction Networks: Analyze potential interactions in the context of known fission yeast protein networks, particularly those involving proteins with similar cell cycle expression patterns or functional domains. The comprehensive cell cycle gene dataset provides valuable context, as genes with similar expression patterns often encode interacting proteins .

• Genetic Interaction Analysis: Combine genetic approaches (synthetic lethality screens, suppressor screens) with protein-level analysis using the SPAC1F7.10 Antibody to correlate genetic and physical interactions .

When undertaking interaction studies, researchers should consider the timing of interactions, particularly if SPAC1F7.10 shows cell cycle-regulated expression, as many protein interactions in fission yeast are temporally regulated during the cell cycle .

What are the common technical challenges when using SPAC1F7.10 Antibody and how can they be addressed?

Researchers working with SPAC1F7.10 Antibody may encounter several technical challenges that require systematic troubleshooting:

• Weak or Absent Signal in Western Blotting:

  • Increase antibody concentration incrementally (starting with manufacturer's recommended dilution)

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

  • Optimize protein extraction methods to ensure preservation of epitopes

  • Verify sample integrity through detection of control proteins

  • Ensure transfer efficiency with Ponceau S staining before immunoblotting

• High Background or Non-specific Bands:

  • Increase blocking time and concentration (5% BSA or milk)

  • Use more stringent washing conditions (increase wash time and buffer volume)

  • Pre-adsorb antibody with acetone powder from negative control samples

  • Titrate antibody to find optimal concentration

  • Include controls to identify non-specific binding patterns

• Variability Between Experiments:

  • Standardize lysate preparation protocols

  • Maintain consistent antibody aliquots to minimize freeze-thaw cycles

  • Implement quantitative loading controls

  • Normalize results to total protein using stain-free technology or Ponceau S

  • Perform technical replicates across multiple protein preparations

• Degraded Antibody:

  • Store according to manufacturer recommendations (-20°C or -80°C)

  • Add sodium azide (0.02%) to diluted antibody solutions for extended use

  • Prepare working aliquots to avoid repeated freeze-thaw cycles

  • Check antibody functionality using positive control samples before critical experiments

• Cross-Reactivity Issues:

  • Perform peptide competition assays to confirm specificity

  • Include appropriate negative controls (knockout strains if available)

  • Optimize antibody concentration to minimize non-specific binding

  • Consider pre-clearing lysates with irrelevant antibodies of the same species

Maintaining detailed records of optimization steps and experimental conditions is crucial for consistent results and troubleshooting.

How can researchers validate lot-to-lot consistency for SPAC1F7.10 Antibody in longitudinal studies?

Ensuring lot-to-lot consistency is crucial for longitudinal studies that may span months or years. Researchers should implement a systematic validation protocol:

• Reference Sample Banking:

  • Prepare and store large batches of positive control samples (wild-type S. pombe lysates)

  • Aliquot and freeze (-80°C) these reference samples to minimize freeze-thaw cycles

  • Use these consistent reference samples to test each new antibody lot

• Comparative Western Blot Analysis:

  • Run side-by-side Western blots using both old and new antibody lots

  • Compare signal intensity, band pattern, and background levels

  • Quantify bands using densitometry to establish quantitative comparison metrics

  • Document acceptable variation thresholds for experimental continuity

• ELISA Calibration Curves:

  • Generate standard curves using recombinant SPAC1F7.10 protein with each antibody lot

  • Compare curve parameters (slope, y-intercept, R² values)

  • Calculate correction factors if necessary to normalize data between lots

• Epitope Verification:

  • Perform peptide competition assays with each new lot

  • Verify that the same epitope is being recognized through consistent competition patterns

• Detailed Record-Keeping:

  • Maintain a database of lot numbers, receipt dates, and validation results

  • Document all optimization parameters for each lot (dilution factors, incubation times)

  • Cross-reference experimental results with specific antibody lots to identify potential lot-related variability

• Long-term Sample Storage:

  • Consider keeping key experimental samples for re-analysis with new antibody lots

  • This approach allows direct comparison of results across the study timeline

Given the made-to-order nature of this antibody (14-16 week lead time), researchers should plan antibody orders well in advance of depletion and consider ordering larger quantities of a single lot for critical longitudinal studies .

What considerations should be made when adapting SPAC1F7.10 Antibody for non-validated applications?

When adapting the SPAC1F7.10 Antibody for applications beyond its validated uses (ELISA and Western blot), researchers should follow a systematic optimization approach:

• Immunofluorescence (IF) Adaptation:

  • Begin with fixation method optimization (test paraformaldehyde, methanol, and acetone fixation)

  • Evaluate multiple permeabilization protocols (Triton X-100, saponin, digitonin at various concentrations)

  • Test a range of antibody concentrations (starting higher than WB dilutions)

  • Include peptide competition controls to verify signal specificity

  • Compare with subcellular markers to validate expected localization patterns

• Immunoprecipitation (IP) Optimization:

  • Test different antibody coupling methods to beads (direct coupling vs. indirect capture)

  • Optimize lysis conditions to preserve protein structure and interactions

  • Evaluate various antibody-to-lysate ratios

  • Include negative controls (non-specific IgG from same species)

  • Confirm pull-down efficiency by Western blot of precipitated fractions

• Chromatin Immunoprecipitation (ChIP) Development:

  • Optimize crosslinking conditions specific to fission yeast

  • Test various sonication/fragmentation parameters

  • Evaluate antibody specificity in the context of crosslinked chromatin

  • Include appropriate controls (IgG, input chromatin)

  • Validate enrichment at expected genomic regions if target has known DNA-binding properties

• Flow Cytometry Adaptation:

  • Develop permeabilization protocols compatible with yeast cell wall

  • Titrate antibody concentration for optimal signal-to-noise ratio

  • Include appropriate controls (unstained, secondary-only, isotype controls)

  • Validate specific staining through competition assays

• Systematic Validation Strategy:

  • Start with positive controls where possible (known localization or interaction partners)

  • Implement genetic controls (overexpression, deletion strains)

  • Document all optimization steps in detail

  • Establish clear criteria for successful adaptation

When adapting this antibody for novel applications, researchers should consider the polyclonal nature of the antibody, which may provide advantages in certain applications (like IP) but might require more extensive validation in others (like IF or ChIP) .

How should researchers interpret variations in SPAC1F7.10 protein levels in the context of fission yeast cell cycle studies?

When interpreting variations in SPAC1F7.10 protein levels throughout the cell cycle, researchers should consider multiple analytical frameworks:

• Peak Expression Timing Analysis:

  • Compare SPAC1F7.10 protein expression patterns to established cell cycle phase markers

  • Utilize the comprehensive cell cycle gene expression dataset as a reference framework, which includes 747 cell cycle-regulated genes with well-defined peak expression times

  • Consider the CDC rank system, where lower numbers indicate stronger cell cycle regulation (e.g., cdc22 with rank 2 shows strong cell cycle-dependent expression)

• Quantitative Temporal Analysis:

  • Apply statistical methods similar to those used in microarray analysis to determine significance of expression changes

  • Consider implementing the modified Fourier-transform method used for gene expression analysis to detect periodicity in protein levels

  • Calculate the False Discovery Rate (FDR) to determine significance of observed variations

• Promoter-Based Interpretation:

  • If SPAC1F7.10 shows cell cycle-regulated expression, analyze its promoter for known regulatory motifs (SFF, MCB, ACE2, ATF)

  • Consider potential transcription factor dependencies based on motif presence

  • Compare with known DSC-dependent or -independent genes based on similar expression patterns

• Comparative Analysis Framework:

  • Evaluate SPAC1F7.10 expression in the context of genes with similar expression patterns

  • Consider potential functional relationships with genes in the same expression cluster

  • Analyze conservation of expression patterns between different synchronization methods (elutriation vs. temperature-sensitive mutants)

• Experimental Variation Assessment:

  • Distinguish between technical variation (antibody performance, sample preparation) and biological variation

  • Implement data normalization strategies similar to those used in gene expression studies

  • Consider potential post-translational modifications that might affect antibody recognition throughout the cell cycle

By applying these analytical frameworks, researchers can generate robust interpretations of SPAC1F7.10 protein expression dynamics in the context of the fission yeast cell cycle.

What approaches can resolve discrepancies between SPAC1F7.10 Antibody results and gene expression data?

Resolving discrepancies between protein-level measurements using SPAC1F7.10 Antibody and corresponding gene expression data requires a multi-faceted approach:

• Temporal Offset Analysis:

  • Plot protein levels against mRNA levels with varying time shifts

  • Calculate correlation coefficients at different offsets to identify optimal lag time

  • Consider the natural delay between transcription and translation (typically 15-30 minutes in yeast)

• Post-transcriptional Regulation Investigation:

  • Analyze mRNA stability using transcriptional inhibitors (e.g., 1,10-phenanthroline)

  • Investigate potential microRNA or RNA-binding protein regulation

  • Examine 3'-UTR sequences for regulatory elements affecting translation efficiency

  • Consider codon optimization analysis that might affect translation rates

• Post-translational Modification Assessment:

  • Evaluate protein stability using cycloheximide chase experiments

  • Investigate potential phosphorylation, ubiquitination, or other modifications

  • Consider whether modifications might affect antibody recognition

  • Implement phosphatase treatments of samples to assess modification impacts

• Technical Validation Approach:

  • Confirm antibody specificity through knockout controls and peptide competition

  • Verify microarray data quality through technical replicates and validation of known markers

  • Consider using alternative detection methods (e.g., epitope tagging) for independent verification

  • Implement spike-in controls for both transcriptomics and proteomics approaches

• Synchronized Culture Optimization:

  • Evaluate synchronization efficiency through morphological examination and flow cytometry

  • Compare results between different synchronization methods (elutriation vs. temperature-sensitive mutants)

  • Consider potential synchronization method-specific artifacts

  • Implement shorter sampling intervals to capture rapid expression changes

When significant discrepancies persist despite these approaches, consider the biological relevance of post-transcriptional regulation as a potential feature of SPAC1F7.10 regulation rather than a technical artifact.

What are the most critical considerations for researchers planning SPAC1F7.10 Antibody-based experiments?

Researchers planning experiments with SPAC1F7.10 Antibody should prioritize several critical considerations to ensure robust and reproducible results:

• Experimental Design Fundamentals:

  • Include appropriate positive and negative controls in every experiment

  • Implement biological and technical replicates to ensure statistical validity

  • Consider temporal aspects, particularly for cell cycle studies, with appropriate synchronization methods

  • Design experiments that account for the polyclonal nature of the antibody and its specific reactivity to S. pombe strain 972/ATCC 24843

• Antibody Handling and Storage:

  • Store at recommended temperatures (-20°C or -80°C)

  • Avoid repeated freeze-thaw cycles by preparing working aliquots

  • Monitor antibody performance over time using consistent positive controls

  • Consider the 14-16 week lead time when planning longitudinal studies

• Application-Specific Optimization:

  • Start with validated applications (ELISA, Western blot) before attempting adaptation

  • Document all optimization steps systematically

  • Establish clear criteria for successful results before proceeding to actual experiments

  • Consider the storage buffer composition (50% glycerol, 0.03% Proclin 300, 0.01M PBS, pH 7.4) when designing compatibility with experimental protocols

• Data Integration Strategies:

  • Plan for integration with existing datasets, particularly gene expression data

  • Consider computational approaches similar to those used in transcriptomic studies

  • Implement appropriate statistical methods for data analysis

  • Design experiments that allow for direct comparison with published cell cycle regulation studies

• Resource and Timeline Planning:

  • Account for the made-to-order nature and extended lead time (14-16 weeks)

  • Consider ordering larger quantities to ensure lot consistency for extended studies

  • Budget for validation experiments before proceeding to critical research applications

  • Plan for potential troubleshooting time in experimental timelines

By carefully considering these critical factors, researchers can maximize the reliability and impact of their SPAC1F7.10 Antibody-based experiments while avoiding common pitfalls that might compromise research outcomes.

How might future research applications expand the utility of SPAC1F7.10 Antibody in fission yeast studies?

Future research applications could significantly expand the utility of SPAC1F7.10 Antibody, opening new avenues for fission yeast studies:

• Multi-omics Integration:

  • Combining antibody-based protein detection with transcriptomics and metabolomics

  • Developing computational models that integrate protein-level data with genome-wide datasets

  • Creating comprehensive regulatory networks incorporating SPAC1F7.10 functionality

  • Applying systems biology approaches similar to those used in cell cycle studies

• Advanced Microscopy Applications:

  • Optimizing the antibody for super-resolution microscopy techniques

  • Developing live-cell imaging approaches using antibody fragments

  • Implementing correlative light and electron microscopy (CLEM) to link protein localization with ultrastructural features

  • Applying single-molecule detection methods to study protein dynamics in living cells

• Evolutionary Conservation Studies:

  • Examining cross-reactivity with homologous proteins in related yeasts

  • Investigating functional conservation through comparative studies

  • Developing antibodies against orthologous proteins in other model organisms

  • Creating evolutionary models of protein function based on cross-species analyses

• Translational Research Applications:

  • Exploring potential connections between SPAC1F7.10 function and human disease models

  • Investigating the role of human homologs in cellular processes and pathologies

  • Developing therapeutic strategies based on conserved mechanisms

  • Implementing fission yeast as a platform for drug screening targeting related pathways

• Technological Innovations:

  • Adapting the antibody for microfluidic and lab-on-a-chip applications

  • Developing biosensor applications using antibody-based detection systems

  • Creating multiplexed detection systems for simultaneous analysis of multiple proteins

  • Implementing AI-driven image analysis for automated protein localization studies