MPZL1 Antibody

What is MPZL1 and what cellular functions does it regulate?

MPZL1 is a transmembrane glycoprotein involved in extracellular matrix-induced signal transduction. It contains an extracellular immunoglobulin-like domain, a transmembrane region, and a cytoplasmic domain with signaling motifs. Research has demonstrated that MPZL1:

• Promotes cell migration and invasion in multiple cancer types

• Activates signaling pathways involving Src kinases and cortactin

• Contributes to cytoskeletal reorganization during cell movement

• Enhances cell proliferation and survival

• May regulate anti-apoptotic responses

MPZL1 is genomically located at chromosome 1q24 and its amplification correlates with increased expression in various cancer types . The protein is expressed at low levels in normal tissues but shows significant upregulation in malignant tissues, particularly in advanced cancers .

What are the common applications for MPZL1 antibodies in research?

MPZL1 antibodies can be employed in several research techniques:

• Western blotting (WB): For detecting protein expression levels and evaluating molecular weight variants. Multiple MPZL1 bands may be observed due to its alternatively spliced isoforms

• Immunohistochemistry (IHC): For examining tissue expression patterns and subcellular localization

• Immunofluorescence (IF): For detailed subcellular localization studies

• Immunoprecipitation (IP): For investigating protein-protein interactions

• ELISA: For quantitative analysis of MPZL1 in biological samples

These applications enable comprehensive investigation of MPZL1's expression, regulation, and functional roles in both normal physiology and disease states.

How do I select the appropriate MPZL1 antibody for my experiments?

When selecting an MPZL1 antibody, consider the following criteria:

For example, when studying MPZL1 in cancer tissues, select antibodies validated for IHC in similar cancer types with demonstrated specificity . Consider antibodies targeting different epitopes (e.g., AA 1-269, AA 107-156, AA 195-258) depending on your research focus .

What controls should I use when working with MPZL1 antibodies?

Implement appropriate controls to ensure reliable results:

• Positive controls: Cell lines or tissues with known MPZL1 expression (e.g., HO8910, HEY, or GBC-SD cells)

• Negative controls: MPZL1 knockdown cells using validated siRNA or shRNA (e.g., siRNA sequences 5′-GCACCUAUAUCUGUGAUGUTT-3′)

• Technical controls: Omit primary antibody or use isotype controls

• Loading controls: Include housekeeping proteins like GAPDH for Western blots

• Antibody validation: Compare results using antibodies targeting different MPZL1 epitopes

For immunohistochemistry, include normal tissues with low MPZL1 expression alongside cancer tissues with expected high expression to establish staining specificity .

What is known about MPZL1 expression in cancer versus normal tissues?

Research demonstrates distinct MPZL1 expression patterns:

• Normal tissues: MPZL1 is expressed at low levels in most normal tissues, including normal ovarian and gallbladder epithelium

• Cancer tissues: Significantly higher expression is observed in malignant tissues compared to benign/borderline tissues

• Cancer progression: Expression correlates with clinical staging, with higher levels in advanced stages (Stage III/IV vs. Stage I/II)

• Metastatic potential: MPZL1 expression positively correlates with metastatic features and invasive capabilities

• Clinical markers: MPZL1 expression correlates with other clinical markers like CA125 levels in ovarian cancer

Immunohistochemical studies have shown that MPZL1 is almost undetectable in benign tissues but shows progressively increasing expression in borderline and malignant conditions .

How can I optimize detection of low MPZL1 expression levels in experimental systems?

For enhanced detection sensitivity:

• Antibody optimization:

  • Test concentration gradients (typically 1:200-1:2000 dilutions)

  • Extend incubation times to overnight at 4°C

  • Compare different antibody clones targeting various epitopes

• Sample preparation:

  • For IHC: Optimize antigen retrieval methods (citrate buffer pH 6.0 or EDTA buffer)

  • For Western blot: Use RIPA buffer with protease inhibitors and phosphatase inhibitors

  • For difficult samples: Consider tissue-specific extraction protocols

• Signal amplification:

  • Implement tyramide signal amplification for IHC/IF

  • Use high-sensitivity ECL reagents for Western blots

  • Apply polymer-based detection systems for increased sensitivity in IHC

• Protein concentration:

  • Increase loading amounts

  • Use immunoprecipitation to concentrate before detection

  • Apply gradient gels for better resolution of isoforms

Published research has successfully detected MPZL1 in various tissues using optimized conditions, including primary antibody concentrations of 1:200 and overnight incubations at 4°C .

How do different experimental approaches affect MPZL1 functional studies?

Various approaches reveal different aspects of MPZL1 function:

Research has demonstrated that MPZL1 knockdown significantly reduces cancer cell migration, invasion, and proliferation, while overexpression enhances these processes .

How can I investigate the relationship between MPZL1 and specific signaling pathways?

To dissect MPZL1's signaling role:

• Pathway component analysis:

  • Examine Src family kinase activation (phospho-Src)

  • Analyze cortactin phosphorylation status

  • Evaluate downstream targets like p130

• Protein-protein interaction studies:

  • Perform co-immunoprecipitation to identify binding partners

  • Use proximity ligation assays for in situ interaction detection

  • Apply crosslinking approaches before mass spectrometry analysis

• Signaling inhibitor treatments:

  • Apply pathway-specific inhibitors to determine dependence

  • Combine with MPZL1 manipulation to establish epistatic relationships

  • Monitor temporal dynamics of pathway activation

• Mutational analyses:

  • Create phosphorylation site mutants to determine critical residues

  • Generate domain deletion constructs to map interaction regions

  • Examine isoform-specific signaling capabilities

Research has established that MPZL1 promotes tumor cell proliferation and migration via activation of specific signaling cascades, including Src-dependent pathways .

What methods are effective for quantifying MPZL1 expression across different cancer stages?

For rigorous quantitative analysis:

• Immunohistochemical scoring:

  • Use standardized scoring systems (percentage of positive cells, staining intensity)

  • Implement blinded evaluation by multiple pathologists

  • Classify expression as negative (-), weak (+), moderate (++), or strong (+++)

• Tissue microarray analysis:

  • Analyze multiple patient samples simultaneously

  • Correlate with clinical parameters (tumor stage, patient age, biomarkers)

  • Document statistically significant associations

• Digital pathology:

  • Apply image analysis software for unbiased quantification

  • Measure both staining intensity and distribution

  • Generate reproducible numerical data

• Molecular quantification:

  • Perform RT-qPCR for transcript level analysis

  • Use Western blot densitometry with appropriate normalization

  • Apply ELISA for standardized protein quantification

Published studies have successfully correlated MPZL1 expression with clinical parameters using these approaches, finding significant associations with tumor stage (p=0.01 for Stage III vs. I-II; p=0.02 for Stage IV vs. III) and biomarkers like CA125 (p=0.01) .

How can I distinguish between the effects of different MPZL1 isoforms?

To investigate isoform-specific functions:

• Molecular identification:

  • MPZL1 has at least three alternatively spliced isoforms visible on Western blots

  • Use gradient gels (4-15% SDS-PAGE) for optimal separation

  • Compare observed bands with predicted molecular weights

• Isoform-specific tools:

  • Design isoform-specific siRNAs/shRNAs

  • Generate expression constructs for individual isoforms

  • Use antibodies recognizing isoform-specific epitopes

• Functional comparison:

  • Perform rescue experiments with specific isoforms after knockdown

  • Compare subcellular localization patterns

  • Examine differential interaction partners

• Expression pattern analysis:

  • Determine tissue-specific isoform distribution

  • Correlate specific isoforms with disease progression

  • Identify conditions that alter isoform ratios

Understanding isoform-specific functions is critical as research indicates that different MPZL1 variants may have distinct biological roles in normal and pathological conditions .

What are optimal protocols for MPZL1 antibody use in FFPE tissue samples?

For effective FFPE tissue analysis:

• Sample preparation:

  • Section thickness: 5 μm sections are optimal

  • Deparaffinization: Complete removal of paraffin is critical

• Antigen retrieval:

  • Heat-induced epitope retrieval in citrate buffer (pH 6.0) for 30 minutes

  • Alternative: EDTA buffer (pH 9.0) for certain epitopes

• Background reduction:

  • Treat with 3% H₂O₂ for 10 minutes to block endogenous peroxidase

  • Use proper blocking solutions (e.g., normal serum matching secondary antibody species)

• Antibody incubation:

  • Primary antibody concentration: 1:200 dilution is commonly effective

  • Incubation time: Overnight at 4°C

  • Secondary antibody: Typically 1:100 dilution for 30 minutes at room temperature

• Detection systems:

  • DAB system for chromogenic detection

  • Counterstain with hematoxylin (5%) for 1 minute

  • Evaluate staining using established scoring criteria

Published protocols have successfully detected MPZL1 in various cancer tissues using these parameters .

How can I effectively troubleshoot inconsistent results with MPZL1 antibodies?

Systematic troubleshooting approaches:

• Antibody validation:

  • Confirm specificity using positive controls (cancer cell lines) and negative controls (siRNA-treated samples)

  • Test antibody lot-to-lot variation against reference samples

  • Consider testing multiple antibodies targeting different epitopes

• Sample quality assessment:

  • Evaluate protein degradation in lysates

  • Check fixation quality in tissue samples

  • Standardize sample collection and processing protocols

• Technical optimization:

  • Titrate antibody concentrations

  • Modify blocking conditions (BSA, milk, commercial blockers)

  • Adjust incubation times and temperatures

  • Try different detection systems

• Controls implementation:

  • Include positive and negative controls in every experiment

  • Use loading controls for Western blots

  • Apply standardized scoring systems for IHC

Comparative analysis of multiple experimental approaches can help reconcile inconsistent results, as demonstrated in studies showing correlation between protein levels, mRNA expression, and functional effects of MPZL1 .

How can MPZL1 antibodies be used to investigate metastatic mechanisms?

To explore MPZL1's role in metastasis:

• Migration and invasion assays:

  • Transwell migration assays show clear MPZL1-dependent effects

  • Compare results before and after MPZL1 manipulation

  • Quantify cell numbers and migration distances

• Metastasis-related pathway analysis:

  • Investigate EMT markers (E-cadherin, vimentin, etc.)

  • Examine matrix metalloproteinase expression/activity

  • Assess cytoskeletal reorganization through cortactin phosphorylation

• In vivo models:

  • Use MPZL1 knockdown or overexpression in xenograft models

  • Quantify metastatic burden in distant organs

  • Correlate MPZL1 expression with metastatic potential

• Clinical sample analysis:

  • Compare MPZL1 expression between primary tumors and metastatic lesions

  • Correlate expression levels with patient outcomes

  • Develop prognostic scoring systems based on MPZL1 status

Research has established that MPZL1 promotes tumor metastasis in multiple cancer types, with mechanistic insights into its activation of pro-metastatic proteins and pathways .