zswim7 Antibody

What is ZSWIM7 and what is its biological significance?

ZSWIM7, also known as SWS1 or zinc finger SWIM domain-containing protein 7, is a nuclear protein that plays a crucial role in the early stages of the homologous recombination repair (HRR) pathway of double-stranded DNA breaks arising during DNA replication or induced by DNA-damaging agents. Research has demonstrated that ZSWIM7 is required for meiotic progression and fertility . The protein forms part of the highly conserved human Shu complex by interacting with SWSAP1, which regulates the recruitment of the strand-exchange protein RAD51 and its homolog DMC1 to meiotic intermediates during homologous recombination .

What types of ZSWIM7 antibodies are available for research?

Multiple types of ZSWIM7 antibodies are available for research applications:

How is ZSWIM7 protein structurally organized?

ZSWIM7 contains a zinc finger SWIM domain that is critical for its function. Mutations affecting this domain, such as the stop-gain mutation p.(Ser58*) reported in patients with Primary Ovarian Insufficiency, likely result in nonsense-mediated decay or protein truncation with loss of this critical domain . For antibody development, specific epitope regions have been targeted, including the amino acid range 68-118 of human ZSWIM7 .

What are the optimal conditions for using ZSWIM7 antibodies in immunohistochemistry?

For immunohistochemistry applications using paraffin-embedded samples, ZSWIM7 antibodies are typically used at dilutions of 1:500-1:1000 . The antibodies should be stored at 4°C for short-term use, while for long-term storage, it is recommended to aliquot and store at -20°C, avoiding freeze-thaw cycles . For optimal results, antigen retrieval methods appropriate for the specific tissue type should be employed before antibody incubation, though specific retrieval conditions vary by antibody manufacturer and should be optimized for each experimental setup.

How should Western blot protocols be optimized for ZSWIM7 detection?

For Western blot applications, ZSWIM7 antibodies are typically used at dilutions ranging from 1:200-1:2000 . When analyzing ZSWIM7 expression by Western blot, researchers should note that transfected ZSWIM7 appears at approximately 15.4 kDa . For optimal results:

• Use protein G purified antibodies for greater specificity

• Employ appropriate blocking solutions (typically 5% non-fat milk or BSA)

• Consider longer primary antibody incubation times (overnight at 4°C) for enhanced sensitivity

• Include appropriate positive controls, such as ZSWIM7-transfected cell lysates compared to non-transfected lysates

What experimental approaches can be used to study ZSWIM7 expression in human reproductive tissues?

Based on research methodologies described in the scientific literature, the following approaches are effective:

• Quantitative RT-PCR: Using TaqMan assays (e.g., ZSWIM7: Hs04984973_m1) with GAPDH (Hs02786624_g1) as a housekeeping control. For human gonadal tissue, samples at critical developmental stages (CS22/23 [7.5-8 wpc], 9 wpc, 11 wpc, 15-16 wpc, and 19-20 wpc) can provide insights into expression during sex differentiation, germ cell expansion, and meiotic entry .

• Immunohistochemistry: Fixed tissue sections from ovary and testis samples can be analyzed using ZSWIM7 antibodies at 1:500-1:1000 dilutions to visualize protein localization patterns .

• RNA sequencing: For comprehensive expression profiling, RNA-seq can be used to compare ZSWIM7 expression with other homologous recombination genes in reproductive tissues .

How can ZSWIM7 antibodies be used to investigate meiotic defects in fertility disorders?

ZSWIM7 antibodies can be valuable tools in investigating meiotic defects in fertility disorders, particularly in cases of Primary Ovarian Insufficiency (POI) or male infertility. Methodological approaches include:

• Immunofluorescence co-localization studies with other meiotic proteins (RAD51, DMC1) in gonadal tissue sections to assess recruitment patterns

• Chromatin immunoprecipitation (ChIP) assays to analyze ZSWIM7 binding to chromatin during meiosis

• Proximity ligation assays (PLA) to detect ZSWIM7 interactions with other components of the Shu complex

Research has shown that ZSWIM7 expression is higher in the 15-16 wpc ovary compared to testis, corresponding with the peak onset of meiosis in the fetal ovary . This temporal expression pattern makes ZSWIM7 antibodies particularly useful for studying early meiotic events in human development.

What approaches can be used to analyze ZSWIM7's role in homologous recombination repair pathways?

To analyze ZSWIM7's role in homologous recombination repair pathways, researchers can employ:

• DNA damage response assays: Treat cells with DNA-damaging agents followed by immunofluorescence with ZSWIM7 antibodies to track protein localization to damage sites

• Co-immunoprecipitation experiments: Use ZSWIM7 antibodies to pull down protein complexes and identify interaction partners in the homologous recombination repair pathway

• CRISPR-Cas9 knockout models: Generate ZSWIM7-deficient cell lines and assess repair efficiency using reporter assays

• Super-resolution microscopy: Examine the spatial organization of ZSWIM7 at sites of DNA damage using antibody-based detection methods

Since ZSWIM7 interacts with Rad51D and plays a role in the early stages of the homologous recombination repair pathway , these approaches can provide insights into the molecular mechanisms underlying DNA repair defects in cells lacking functional ZSWIM7.

How does ZSWIM7 expression change during meiotic progression, and how can this be quantified?

Research has demonstrated distinct temporal expression patterns of ZSWIM7 during meiotic progression. To quantify these changes:

• Comparative qRT-PCR analysis: ZSWIM7 expression shows higher levels in the 15-16 wpc ovary (peak meiosis onset) compared to testis, while adult testis (with ongoing meiosis) shows relatively strong expression . Researchers can use qRT-PCR with appropriate housekeeping genes to quantify these changes across developmental timepoints.

• Protein expression quantification: Western blot analysis using calibrated ZSWIM7 antibodies can provide protein-level quantification during different meiotic stages.

• Single-cell RNA sequencing: For more granular analysis, scRNA-seq can identify cell populations with varying ZSWIM7 expression levels during gametogenesis.

The expression profile of ZSWIM7 correlates with known meiotic gene expression patterns, suggesting its coordinated regulation with other homologous recombination factors during prophase I .

How can researchers verify ZSWIM7 antibody specificity in their experimental systems?

To verify ZSWIM7 antibody specificity, researchers should:

• Include positive controls: Use ZSWIM7-transfected cell lysates alongside non-transfected controls for Western blot analysis

• Validate with knockout/knockdown controls: Compare antibody signal in wild-type versus ZSWIM7-depleted samples

• Peptide competition assays: Pre-incubate antibodies with blocking peptides (e.g., sc-136956 P ) to confirm signal specificity

• Cross-validation with multiple antibodies: Use different antibodies targeting distinct epitopes of ZSWIM7 to confirm consistent detection patterns

• Verify by mass spectrometry: Confirm identity of immunoprecipitated proteins when using ZSWIM7 antibodies for pull-down experiments

Some commercially available ZSWIM7 antibodies have been verified on protein arrays containing target protein plus 383 other non-specific proteins to ensure specificity .

What methodological approaches should be used when interpreting contradictory ZSWIM7 expression data?

When faced with contradictory ZSWIM7 expression data, researchers should:

• Consider tissue/cell type differences: ZSWIM7 expression varies across tissues, with notable expression in reproductive tissues but also ubiquitous expression elsewhere, suggesting involvement in recombinational DNA repair pathways outside of meiosis

• Evaluate developmental timing: Expression patterns change significantly during development, particularly in gonadal tissues, making temporal considerations crucial

• Compare detection methods: Cross-validate findings using multiple techniques:

  • qRT-PCR for mRNA levels

  • Western blot for protein expression

  • Immunohistochemistry for spatial localization

• Examine splice variants: Check if contradictory data might be due to detection of different ZSWIM7 isoforms

• Reference public databases: Compare findings to data from GTEx, Human Protein Atlas, and FANTOM5 to contextualize results within broader expression patterns

How can researchers optimize ZSWIM7 antibody-based detection in tissues with low expression levels?

For detecting ZSWIM7 in tissues with low expression levels, researchers should consider:

• Signal amplification methods:

  • Tyramide signal amplification (TSA) for immunohistochemistry

  • Enhanced chemiluminescence (ECL) substrates with extended exposure times for Western blot

  • Biotin-streptavidin amplification systems

• Sample enrichment techniques:

  • Immunoprecipitation before Western blot analysis

  • Laser capture microdissection to isolate specific cell populations from tissue sections

• Optimized blocking conditions:

  • Test different blocking agents (BSA, non-fat milk, normal serum)

  • Extend blocking times to reduce background

• Extended antibody incubation:

  • Longer primary antibody incubation (overnight at 4°C)

  • Optimized antibody concentration through titration experiments

• Sensitive detection systems:

  • Fluorescent secondary antibodies with appropriate filters

  • Confocal microscopy for improved signal-to-noise ratio

How can ZSWIM7 antibodies be used to investigate potential therapeutic approaches for fertility disorders?

Since ZSWIM7 mutations have been associated with Primary Ovarian Insufficiency , antibodies against this protein could be valuable tools for investigating potential therapeutic approaches:

• Screening potential drug candidates that modulate homologous recombination repair pathways

• Monitoring restoration of proper meiotic progression in experimental models

• Developing diagnostic approaches to identify patients with ZSWIM7-related fertility disorders

• Investigating timing of therapeutic interventions, particularly in males where meiosis occurs post-puberty

In knockout mouse models, disruption of ZSWIM7/SWS1 leads to fertility defects similar to human POI, making these research tools valuable for translational studies .

What methodological considerations are important when using ZSWIM7 antibodies in single-cell analysis techniques?

For single-cell applications using ZSWIM7 antibodies, researchers should consider:

• Antibody validation: Confirm specificity in single-cell contexts where target protein may be present at very low abundance

• Fixation optimization: Test multiple fixation protocols to preserve epitope accessibility while maintaining cellular morphology

• Signal amplification: Implement appropriate amplification techniques compatible with single-cell resolution

• Multiplexing strategies: Combine ZSWIM7 antibodies with other markers to identify specific cell populations or co-localization with interaction partners

• Image analysis: Utilize appropriate quantification algorithms to measure signal intensity and subcellular localization at the single-cell level