mboa-7 Antibody

Basic Research Questions

How should researchers validate mboa-7 antibody specificity in Western blot experiments?

• Methodological approach:

  • Use knockout controls (e.g., CRISPR-edited MBOAT7-deficient cell lines or tissues) to confirm absence of signal at 45 kDa .

  • Compare observed vs. calculated molecular weights (45 kDa observed vs. 53 kDa predicted) to identify potential isoforms or post-translational modifications .

  • Perform peptide competition assays using immunogen-derived peptides to confirm epitope specificity .

What experimental conditions optimize mboa-7 detection in mouse liver tissue?

• Key parameters:

  Parameter	Recommendation
  Primary antibody dilution	1:500–1:1000
  Blocking buffer	5% non-fat milk in TBST
  Secondary antibody	HRP-conjugated anti-rabbit IgG (1:5,000)
  Detection method	Enhanced chemiluminescence (ECL)
  Reference: Proteintech protocol for 26646-1-AP .

How does mboa-7 antibody facilitate studies on phosphatidylinositol (PI) remodeling?

• Functional context:

  • Use the antibody to track MBOAT7 localization in ER/Golgi fractions via subcellular fractionation .

  • Pair with lipidomic profiling to correlate protein expression levels with arachidonic acid (AA) incorporation into PI .

Advanced Research Questions

How to resolve discrepancies between predicted and observed molecular weights of MBOAT7?

• Hypothesis-driven troubleshooting:

  • Potential causes: Alternative splicing (e.g., UniProt Q96N66 lists multiple isoforms) or proteolytic cleavage .

  • Validation steps:

    • Perform RT-PCR to identify splice variants in your model system.

    • Use protease inhibitors (e.g., PMSF) during protein extraction to prevent degradation .

What strategies address low signal-to-noise ratios in co-localization studies with ACSL4/MBOAT7?

• Experimental design:

  Approach	Rationale
  Sequential staining	Minimize cross-reactivity between primary antibodies
  Super-resolution microscopy	Resolve ER/Golgi membrane proximity (≤200 nm)
  Proximity ligation assays	Confirm physical interaction (e.g., MMD scaffold role)

How to design functional assays linking MBOAT7 to ferroptosis susceptibility?

• Integrated workflow:

  • Knockdown MBOAT7 in ovarian carcinoma cells using siRNA .

  • Measure lipid peroxidation via C11-BODIPY staining.

  • Quantify AA-PI levels using LC-MS/MS .

  • Critical controls: Include ACSL4-deficient cells to dissect metabolic flux contributions .

Data Interpretation & Contradictions

How to reconcile conflicting reports on MBOAT7’s role in neuronal vs. cancer biology?

• Comparative analysis framework:

  Biological context	MBOAT7 function	Key readouts
  Neuronal development	AA-PI synthesis defects	Intellectual disability, epilepsy
  Cancer (ovarian/renal)	Ferroptosis regulation	Lipid peroxidation, ACSL4 interaction

  • Solution: Use tissue-specific knockout models to isolate context-dependent roles.

What molecular mechanisms explain reduced LPIAT activity in mboa-7 mutants?

• Mechanistic validation:

  • Express recombinant human MBOAT7 in C. elegans mutants to rescue EPA-PI levels .

  • Perform in vitro acyltransferase assays with AA-CoA and lyso-PI substrates .

  • Key data: AA-CoA incorporation rates drop by ≥70% in mutants .

Methodological Innovations

Can mboa-7 antibody be adapted for single-cell resolution studies?

• Protocol adaptation:

  • Combine with PLA (Proximity Ligation Assay) to quantify MBOAT7/ACSL4 complexes in fixed cells .

  • Use antibody-conjugated quantum dots for multiplexed imaging in live-cell systems.

How to standardize cross-species reactivity in comparative studies?

• Validation matrix:

  Species	Reactivity	Validation method
  Human	Confirmed (WB/IF)	HEK293T lysates
  Mouse	Confirmed (WB)	Liver tissue
  C. elegans	Indirect (functional rescue)	Transgenic mboa-7 mutants