oig-1 Antibody

FAQs for OIG-1 Antibody Research
Below are structured research-focused questions addressing key methodological and conceptual challenges in studying the C. elegans OIG-1 protein, synthesized from peer-reviewed studies.

How to detect endogenous OIG-1 protein expression in GABA neurons?

• Methodology:

  • Use transgenic strains expressing fluorescent reporters (e.g., Poig-1::GFP or mCherry::OIG-1 fusions) under GABA-specific promoters like unc-25 .

  • Validate via live antibody labeling (e.g., anti-OIG-1 antibodies) combined with immunofluorescence in oig-1(ok1687) mutants to confirm specificity .

  • Critical controls: Include coelomocyte staining to confirm secretion (full-length OIG-1) vs. intracellular retention (OIG-1-SP, lacking signal peptide) .

What experimental designs validate OIG-1’s role in synaptic remodeling timing?

• Approach:

  • Compare ACR-12::GFP puncta dynamics in wild-type vs. oig-1 null mutants using time-lapse imaging (8–18 hours post-hatching) .

  • Rescue experiments: Express OIG-1 variants (secreted vs. non-secreted) in GABA neurons (unc-25 promoter) or cholinergic neurons (unc-17 promoter) to test cell autonomy .

  • Key metric: Quantify dorsal-to-ventral ACR-12::GFP redistribution rates (Figure 1F in ).

Does OIG-1 stabilize ACR-12 receptor clusters intracellularly or extracellularly?

• Resolution:

  • Intracellular role: Non-secreted OIG-1-SP rescues dorsal ACR-12::GFP puncta in mutants, confirmed by absent coelomocyte uptake .

  • Extracellular role: Full-length OIG-1 secretion is dispensable for remodeling, as shown by mosaic expression analysis (Figure 4B–C in ).

How to resolve conflicting data on OIG-1’s secretion versus intracellular function?

• Contradiction: While OIG-1 is secreted (evidenced by coelomocyte uptake), its synaptic stabilization does not require secretion .

• Experimental strategies:

  • Compare functional rescue efficiency between full-length OIG-1 and OIG-1-SP using Pflp-13::ACR-12::GFP reporters.

  • Perform surface vs. intracellular staining with pH-sensitive antibodies to distinguish membrane-bound vs. internalized pools .

  • Key finding: OIG-1-SP restores dorsal ACR-12::GFP clusters despite lacking secretion (Figure 4C in ).

What molecular mechanisms regulate OIG-1 downregulation in DD neurons during L1/L2 transition?

• Hypothesis testing:

  • Screen transcriptional regulators of oig-1 using GFP reporters with truncated promoters (e.g., Poig-1Δ300) .

  • CRISPR-Cas9-mediated deletion of candidate transcription factor binding sites in the oig-1 promoter.

  • Evidence: oig-1 expression ceases in DD neurons by L2, correlating with dorsal synapse formation .

How to integrate OIG-1 studies with other synaptic organizers (e.g., neuroligins, Wnts)?

• Multiplexed approach:

  • Use triple-transgenic strains (e.g., oig-1 mutants + nlp-12 mutants) to assess redundant pathways.

  • Map spatial overlap of OIG-1::mCherry with Wnt receptor LIN-17::GFP using super-resolution microscopy.

  • Table: Known synaptic organizers in C. elegans DD remodeling:

Methodological Best Practices

• Antibody validation:

  • Use oig-1(ok1687) null mutants as negative controls for immunostaining .

  • For western blotting, combine anti-OIG-1 antibodies with fluorescent secondaries (e.g., Alexa Fluor 488 goat anti-mouse) to enhance signal-to-noise .

• Quantitative analysis:

  • Calculate enrichment ratios for ACR-12::GFP puncta using Fiji/ImageJ macros (e.g., particle analysis with size thresholds) .