At1g22430 Antibody

Basic Research Questions

What is the molecular function of At1g22430, and how does its antibody aid in characterization?

At1g22430 encodes a GroES-like zinc-binding dehydrogenase family protein involved in redox metabolism and potential stress responses . Its antibody enables:

• Subcellular localization via immunofluorescence or immunogold electron microscopy

• Expression profiling under abiotic/biotic stress using western blot or ELISA

• Protein-protein interaction studies (e.g., co-immunoprecipitation with metabolic enzymes)

What validation methods are critical for ensuring At1g22430 antibody specificity?

How can this antibody be applied in mutant phenotyping studies?

• Compare protein abundance in T-DNA insertion lines (e.g., SALK_XXXXXX) vs wild-type

• Monitor compensatory mechanisms in RNAi knockdowns under hypoxia

• Validate CRISPR-Cas9 editing efficiency via loss-of-signal assays

Advanced Research Questions

How to resolve contradictions in tissue-specific expression data?

Conflicting reports may arise from:

• Post-translational modifications altering epitope accessibility

• Cross-reactivity with paralogs (e.g., AT1G22440, a homologous dehydrogenase)

• Fixation artifacts in immunohistochemistry

Methodological solutions:

• Perform 2D gel electrophoresis to detect PTM isoforms

• Use knockout-validated multiple reaction monitoring (MRM) mass spectrometry

• Optimize antigen retrieval protocols for plant tissues

What experimental designs mitigate off-target effects in live-cell imaging?

• Dual-labeling: Co-stain with organelle markers (e.g., mitochondrial COXII)

• Fluorescence lifetime imaging (FLIM) to distinguish autofluorescence

• Transient expression: Fuse At1g22430 with GFP in protoplasts as internal control

How to integrate At1g22430 antibody data with multi-omics datasets?

What mechanisms explain discordance between antibody reactivity and enzyme activity assays?

• Allosteric regulation: Antibody binding may stabilize inactive conformations

• Competitive inhibitors in crude extracts masking epitopes

• Tissue-specific proteolysis generating truncated immunoreactive fragments

Resolution workflow:

• Compare native vs denaturing western blot conditions

• Test antibody reactivity against recombinant protein pre-treated with metabolites

• Quantify activity in immunodepleted extracts

Technical Notes

• Critical controls: Always include Arabidopsis ecotype-matched wild-type and knockout lines

• Storage: Aliquot antibodies in 50% glycerol at -80°C; avoid freeze-thaw cycles

• Ethical reporting: Disclose lot-to-lot variability and validation data in publications