Customize ADF1 Antibody

Description

Definition and Biological Context

ADF1 antibodies are immunoglobulins designed to bind specifically to the ADF1 protein, a member of the actin-depolymerizing factor (ADF)/cofilin family. ADF1 regulates actin filament dynamics by severing and depolymerizing actin, critical for processes such as cell division, motility, and structural reorganization .

Research Applications and Findings

ADF1 antibodies have been pivotal in elucidating mechanisms of cytokinesis and actin regulation across model organisms:

Fission Yeast (Schizosaccharomyces pombe)

Role in Cytokinesis: ADF1 antibodies revealed that Adf1 is essential for contractile ring assembly by depolymerizing actin at cell ends and severing filaments .
Collaborative Interactions: Genetic studies showed Adf1 works with profilin, capping protein, and myosin-II, while tropomyosin antagonizes its activity .

Parasites (Toxoplasma gondii and Plasmodium falciparum)

Functional Conservation: PfADF1 (Plasmodium) complemented TgADF (Toxoplasma) in rescue experiments, highlighting evolutionary conservation .
Mutational Analysis: Lysine residue mutations (e.g., K68A in TgADF) impaired actin disassembly, disrupting organelle segregation but not motility .

Mechanisms of Action

ADF1 antibodies enable visualization and functional disruption of ADF1 activity:

Actin Depolymerization: ADF1 binding accelerates actin filament turnover, critical for mitotic actin reorganization .
Contractile Ring Maintenance: Antibody-based assays demonstrated Adf1’s role in stabilizing the ring structure post-assembly .

Comparative Analysis

ADF1 antibodies differ in utility compared to related antibodies:

Feature	ADF1 Antibody	Anti-Actin Antibody
Primary Target	ADF1 protein	Actin filaments
Function Highlighted	Actin severing/depolymerization	Filament stability and organization
Key Applications	Cytokinesis studies, parasite development	Cell structure imaging, motility assays

Table 1: ADF1 Antibody Applications in Select Studies

Study	Organism	Method	Key Finding
Nakano et al. (2006)	Fission yeast	Immunoblotting	Adf1 required for contractile ring assembly .
Wong et al. (2015)	Toxoplasma gondii	Immunofluorescence	ADF1 Lys-68 mutation disrupts actin disassembly .

Table 2: Biochemical Properties of ADF1

Property	Detail
Actin Severing Rate	~0.0008 breaks/μm/s (in vitro)
Critical Residues	Lys-68 (TgADF), Lys-72 (PfADF1)
Binding Partners	Profilin, myosin-II, tropomyosin

Technical Considerations

Immunofluorescence Protocols: Fixation with paraformaldehyde/glutaraldehyde and Triton X-100 permeabilization are standard for ADF1 localization .
Western Blotting: Anti-Adf1 antibodies (affinity-purified) and anti-HA/anti-His tags are used for detection .

Challenges and Future Directions

Current limitations include genus-specific variations in ADF1 function, necessitating species-specific antibody validation . Advances in cryo-EM and CRISPR-based tagging could enhance spatial resolution of ADF1 dynamics.

Product Specs

Buffer

**Preservative:** 0.03% Proclin 300
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4

Form

Liquid

Lead Time

Made-to-order (14-16 weeks)

Synonyms

ADF1 antibody; At3g46010 antibody; F16L2_220Actin-depolymerizing factor 1 antibody; ADF-1 antibody; AtADF1 antibody

Target Names

ADF1

Uniprot No.

Q39250

Target Background

Function

ADF1 (Actin Depolymerizing Factor 1) is a protein that promotes the disassembly of actin filaments. It plays a crucial role in plant development, cell organ expansion, and flowering by regulating the breakdown of thick actin cables. ADF1 severs actin filaments or bundles, leading to the disassembly of the actin cytoskeleton. It binds to monomeric actin (G-actin) with a strong preference for the ADP-loaded form, inhibiting the rate of nucleotide exchange on G-actin.

Gene References Into Functions

Research indicates that 14-3-3 lambda is essential for the phosphorylation and subcellular localization of ADF1. PMID: 26345162
Studies have demonstrated a regulatory role of Arabidopsis CDPK6 in the N-terminal phosphorylation of AtADF1. PMID: 23903947
Mutations on actin binding sites within Arabidopsis AtADF1 have been shown to affect its function. PMID: 23190411

Database Links

KEGG: ath:AT3G46010

STRING: 3702.AT3G46010.2

UniGene: At.21187

Protein Families

Actin-binding proteins ADF family

Subcellular Location

Cytoplasm, cytoskeleton.

Tissue Specificity

Expressed in vascular tissues of all organs.

Q&A

FAQs for ADF1 Antibody in Academic Research

Advanced Research Questions

How do ADF1 genetic interactions inform its role in actin cytoskeleton dynamics?

Methodological Answer:
Conduct epistasis analysis in S. pombe:

Protein Interaction	Phenotype in Double Mutants	Key Finding
Adf1 + Profilin	Synthetic lethality	Coordinated actin depolymerization and monomer recycling
Adf1 + Myosin-II	Defective contractile ring assembly	Collaboration in ring stabilization
Adf1 + Tropomyosin	Rescue of ring instability	Antagonistic regulatory relationship

How can researchers resolve contradictions in ADF1’s subcellular localization across studies?
- Methodological Answer:
  - Fixation optimization: Use methanol-free formaldehyde to preserve actin structures in fission yeast .
  - Antibody cross-reactivity checks: Test antibodies against Drosophila ADF1 in S. pombe lysates to rule out nonspecific binding .

Methodological Challenges

What controls are essential for co-localization studies using ADF1 antibodies and cytoskeletal markers?
- Methodological Answer:
  - Include secondary antibody-only controls to eliminate autofluorescence.
  - Use dual staining with established markers (e.g., Cdc8 for actin patches, tubulin for microtubules) .
  - Quantify colocalization using Pearson’s correlation coefficient (>0.7 indicates significant overlap) .
How to design epistasis experiments for ADF1’s transcriptional role in Drosophila?
- Methodological Answer:
  - Perform ChIP-seq with anti-Adf1 antibodies in embryos at 4–5 hours post-fertilization to map binding sites near the Adh promoter .
  - Combine nal P1 mutants (reduced ADF1) with Adh reporter constructs to assess transcriptional activation .

Data Analysis & Interpretation

How should researchers interpret conflicting data on ADF1’s role in synaptic bouton formation?
- Methodological Answer:
  - Compare expression levels: nal P1 mutants show reduced boutons, while ADF1 overexpression increases them .
  - Control for neuronal activity: Use electrophysiology (e.g., evoked transmitter release assays) to distinguish structural vs. functional defects .
What statistical approaches are robust for quantifying ADF1-dependent actin depolymerization?
- Methodological Answer:
  - Use fluorescence recovery after photobleaching (FRAP) to measure actin turnover rates in adf1 null vs. wild-type cells .
  - Apply ANOVA with post-hoc Tukey tests for multi-group comparisons (e.g., Adf1 + profilin vs. Adf1 + capping protein mutants) .

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ADF1 Antibody