MRPL42 Antibody

What is MRPL42 and why is it significant for research?

MRPL42 (mitochondrial ribosomal protein L42) is a nuclear-encoded protein that functions in mitochondrial protein synthesis. It belongs to both the 28S and 39S subunits of mitoribosomes, which have an estimated 75% protein to rRNA composition compared to prokaryotic ribosomes (where this ratio is reversed) . Recent research has revealed MRPL42's significance beyond its structural role, as it has been implicated in several cancer types including lung adenocarcinoma and glioma .

The protein has a calculated molecular weight of approximately 17 kDa (142 amino acids) and typically shows observed molecular weight of 14-17 kDa in experimental settings . MRPL42 is also known by several aliases including L31mt, L42mt, MRP-L31, MRP-L42, MRP-S32, MRPL31, MRPS32, PTD007, RPML31, and S32MT .

How do I select the appropriate MRPL42 antibody for my research?

Selection should be guided by:

• Application compatibility: Verify the antibody has been validated for your specific application (WB, IHC, IF/ICC, or ELISA). For example, the Proteintech 17300-1-AP antibody has been validated for WB (1:500-1:3000 dilution), IHC (1:500-1:2000 dilution), and IF/ICC (1:200-1:800 dilution) .

• Species reactivity: Most commercially available MRPL42 antibodies show reactivity with human samples . If working with other species, verify cross-reactivity or consider specialized antibodies.

• Clonality requirements:

  • Polyclonal antibodies (e.g., Proteintech 17300-1-AP, Elabscience E-AB-19123) offer broader epitope recognition

  • Monoclonal antibodies (e.g., Invitrogen MA5-17128 clone 3H6G11, Abcam ab279370 clone OTI1A4) provide higher specificity and reproducibility

• Validation evidence: Review literature and product documentation showing specific detection of MRPL42 in relevant experimental systems .

What are the optimal protocols for using MRPL42 antibodies in cancer tissue samples?

Based on published research and manufacturer recommendations:

For IHC applications:

• Tissue preparation: Use formalin-fixed paraffin-embedded (FFPE) tissues with appropriate antigen retrieval methods:

  • Heat-induced epitope retrieval with TE buffer (pH 9.0) is recommended for most MRPL42 antibodies

  • Alternatively, citrate buffer (pH 6.0) can be used

• Antibody dilution:

  • For polyclonal antibodies: 1:500-1:2000 (Proteintech)

  • For monoclonal antibodies: 1:150 (Abcam ab279370)

• Detection method: Both MRPL42 polyclonal and monoclonal antibodies have been successfully used for detecting MRPL42 in various cancer tissues including breast cancer, thyroid carcinoma, pancreatic tissue, and cervical cancer .

For IF/ICC applications:

• Cell preparation: Demonstrated successful detection in various cell lines including HeLa cells

• Antibody dilution: 1:200-1:800 (Proteintech)

• Visualization: Fluorescent secondary antibodies with appropriate controls

What are the critical factors for successful Western blot detection of MRPL42?

• Sample preparation:

  • Effective for detection in multiple cell lines: HeLa, HepG2, MCF-7, Jurkat, and PC-3 cells

  • Complete lysis buffers containing protease inhibitors are essential

• Loading and transfer parameters:

  • Load 20-40 μg of total protein per lane

  • Use appropriate molecular weight markers (MRPL42 typically observed at 14-17 kDa)

• Antibody concentration:

  • Primary antibody dilution: 1:500-1:3000 for most polyclonal antibodies

  • Secondary antibody selection should match the host species of primary antibody

• Detection considerations:

  • Enhanced chemiluminescence (ECL) systems work well for MRPL42 detection

  • Expected band at 14-17 kDa, consistent with predicted molecular weight

How can I design MRPL42 knockdown experiments to investigate its role in cancer progression?

Based on published studies of MRPL42 knockdown:

• shRNA approach:

  • Lentiviral-mediated shRNA has been successfully used to knockdown MRPL42 in lung adenocarcinoma (A549, H1299) and glioma (U251, A172) cell lines

  • Construct design should target conserved regions of MRPL42 mRNA

  • Include appropriate controls (sh-blank, sh-ctrl) to validate specificity

• Verification methods:

  • Validate knockdown efficiency via qRT-PCR (mRNA level) and Western blot (protein level)

  • Successful knockdowns typically show >70% reduction in MRPL42 expression

• Phenotypic assays:

  • Cell proliferation: CCK-8 assay, colony formation assay, and high-content screening (HCS)

  • Cell cycle analysis: Flow cytometry to assess G1/S phase distribution

  • Apoptosis: Caspase-3/caspase-7 activity assays

  • Migration and invasion: Transwell migration assay, Matrigel invasion assay

• In vivo validation:

  • Nude mice xenograft models have confirmed in vitro findings

  • Compare tumor weight, growth rate, and expression of proliferation markers (Ki67) and metastasis markers (Vimentin) between control and MRPL42-knockdown tumors

What are the molecular mechanisms underlying MRPL42's oncogenic activity?

Current research has identified several mechanisms:

• Transcriptional regulation:

  • MRPL42 is activated by the transcription factor YY1 in lung adenocarcinoma

  • ChIP analysis has confirmed YY1 binding to the MRPL42 promoter

  • siRNA-mediated knockdown of YY1 significantly decreases MRPL42 expression

• Cell cycle regulation:

  • MRPL42 knockdown induces G1/S cell cycle arrest in glioma and lung adenocarcinoma cells

  • This suggests MRPL42 promotes cell cycle progression at the G1/S checkpoint

• Apoptotic pathways:

  • MRPL42 silencing activates apoptosis and increases caspase-3/caspase-7 activity

  • Historical research suggests MRPL42 may interact with Bcl-2 and activate JNK signaling

• Metastasis promotion:

  • MRPL42 expression correlates with lymph node metastasis in LUAD patients

  • Knockdown reduces expression of metastasis markers like Vimentin

  • Clinical data shows correlation between MRPL42 expression and tumor size (p=0.003) and lymph node metastasis (p=0.032) in LUAD patients

How can I minimize background and optimize signal-to-noise ratio when using MRPL42 antibodies?

• For Western blot applications:

  • Increase blocking time (5% BSA or non-fat milk for 1-2 hours)

  • Optimize primary antibody concentration through titration experiments (start with recommended 1:500-1:3000 dilution)

  • Include appropriate washing steps (3-5 washes of 5-10 minutes each)

  • Use freshly prepared buffers and reagents

  • Consider gradient SDS-PAGE to better resolve the 14-17 kDa region

• For IHC applications:

  • Increase blocking time and optimize antibody dilution (1:500-1:2000)

  • Compare antigen retrieval methods (TE buffer pH 9.0 vs. citrate buffer pH 6.0)

  • Include appropriate negative controls (isotype control or secondary antibody only)

  • For high background, consider increasing washing steps or reducing antibody concentration

• For IF/ICC applications:

  • Use 0.1-0.3% Triton X-100 for proper permeabilization

  • Optimize antibody concentration (1:200-1:800 recommended)

  • Include DAPI nuclear counterstain for proper cellular localization evaluation

  • Use multiple channels to confirm specificity and mitochondrial localization

How should I interpret discrepancies in MRPL42 detection between different antibodies or techniques?

• Epitope accessibility variations:

  • Different antibodies target different regions of MRPL42

  • Monoclonal antibodies recognize specific epitopes, while polyclonal antibodies recognize multiple epitopes

  • Some epitopes may be masked in certain experimental conditions or cell types

• Post-translational modifications:

  • MRPL42 may undergo modifications that affect antibody recognition

  • Compare results from multiple antibodies targeting different epitopes

• Validation strategies:

  • Use MRPL42 knockdown controls to confirm specificity

  • Verify with alternative detection methods (e.g., mass spectrometry)

  • Compare results with published literature on MRPL42 expression patterns

• Technical considerations:

  • Sample preparation methods can affect MRPL42 detection

  • Storage conditions of antibodies may impact performance

  • Consider using newly developed antibodies with improved specificity

What is the correlation between MRPL42 expression and clinical parameters in cancer patients?

Data from clinical studies shows significant correlations:

In Lung Adenocarcinoma (LUAD):

*P<0.05 indicates statistical significance

Key findings:

• MRPL42 expression is significantly associated with tumor size (p=0.003)

• MRPL42 expression correlates with lymph node metastasis (p=0.032)

• No significant correlation with gender, age, or smoking status

In Glioma:

• MRPL42 expression is significantly higher in glioma tissues compared to normal tissues according to TCGA database analysis

• Knockdown studies demonstrate its role in glioma cell proliferation and survival

How can MRPL42 antibodies be utilized for developing cancer diagnostic or prognostic tools?

Based on current research findings:

• Diagnostic applications:

  • IHC staining of MRPL42 in tissue biopsies shows potential for distinguishing cancer tissues

  • MRPL42 antibodies have been validated in multiple cancer types including breast cancer, thyroid carcinoma, pancreatic tissue, and cervical cancer

  • Comparative analysis with normal tissues shows significant upregulation in tumors

• Prognostic markers:

  • High MRPL42 expression correlates with poor prognosis in LUAD patients

  • Association with tumor size and lymph node metastasis suggests utility as a prognostic biomarker

  • Combined panels with other mitochondrial ribosomal proteins might enhance prognostic value

• Methodological considerations:

  • Standardized IHC protocols with optimal antibody dilutions (1:500-1:2000)

  • Consistent scoring systems based on staining intensity and distribution

  • Integration with clinical parameters for comprehensive evaluation

• Future directions:

  • Development of MRPL42-targeted therapy based on its oncogenic properties

  • Combination with other biomarkers for improved specificity and sensitivity

  • Exploration of circulating MRPL42 as a non-invasive biomarker

What are the comparative advantages of different methods for generating MRPL42-specific antibodies?

Based on experimental evidence:

• Polyclonal antibodies against MRPL42 fusion proteins have demonstrated good specificity in multiple applications

• Monoclonal antibodies targeting specific MRPL42 epitopes show excellent performance in IHC applications

• The choice depends on research needs: polyclonals for broader epitope recognition, monoclonals for higher specificity

What validation steps are critical for ensuring MRPL42 antibody specificity?

A comprehensive validation strategy includes:

• Expression systems verification:

  • Positive detection in known MRPL42-expressing cell lines (HeLa, HepG2, MCF-7, Jurkat, PC-3)

  • Comparative analysis with normal cell lines (e.g., 16HBE for lung studies)

• Knockdown/knockout controls:

  • shRNA-mediated knockdown models as negative controls

  • Confirm reduction/absence of signal in MRPL42-depleted samples

• Cross-validation with multiple techniques:

  • Compare protein detection by Western blot, IHC, and IF

  • Verify signal correlates with mRNA expression (qRT-PCR)

  • Mass spectrometry confirmation where possible

• Epitope mapping:

  • Identify specific binding regions within MRPL42

  • Validate against recombinant protein fragments

  • For example, MA5-17128 targets amino acids 142-203 of human MRPL42

• Species cross-reactivity assessment:

  • Test antibody performance across relevant species

  • Most MRPL42 antibodies are optimized for human samples

How is MRPL42 being investigated as a potential therapeutic target in cancer?

Current research explores several promising approaches:

• Gene silencing strategies:

  • shRNA and siRNA approaches have demonstrated anti-tumor effects in lung adenocarcinoma and glioma models

  • In vivo studies show reduced tumor growth in nude mice injected with MRPL42-knockdown cells

• Disruption of transcriptional activation:

  • Targeting the YY1-MRPL42 axis may provide therapeutic opportunities

  • ChIP and dual luciferase reporter assays have confirmed YY1 as a transcriptional activator of MRPL42

• Combination therapies:

  • MRPL42 knockdown has been shown to:

    • Reduce p-Akt expression

    • Decrease Vimentin levels

    • Induce G1/S cell cycle arrest

  • These mechanisms could be targeted alongside standard therapies

• Potential challenges:

  • Mitochondrial localization may complicate drug delivery

  • Essential role in normal cells requires careful targeting of cancer-specific pathways

  • Development of MRPL42-specific inhibitors that don't affect other mitoribosomal proteins

What novel techniques are advancing our understanding of MRPL42's function in mitochondrial translation?

Emerging methodologies in this field include:

• Advanced imaging techniques:

  • Super-resolution microscopy to visualize MRPL42 within mitoribosome complexes

  • Live-cell imaging to track MRPL42 dynamics during mitochondrial translation

  • MRPL42 antibodies conjugated to fluorescent tags enable these applications

• Structural biology approaches:

  • Cryo-EM studies of mitoribosome structure with MRPL42

  • Analysis of MRPL42's position within both 28S and 39S subunits

  • Investigation of potential binding partners within the mitoribosome

• Proximity-based proteomics:

  • BioID or APEX2 tagging of MRPL42 to identify interacting proteins

  • Mass spectrometry analysis of MRPL42 complexes in different cellular contexts

  • Comparison between normal and cancer cells to identify cancer-specific interactions

• Single-cell analysis:

  • Single-cell RNA-seq to characterize MRPL42 expression heterogeneity

  • Correlation with mitochondrial activity and cellular phenotypes

  • Integration with spatial transcriptomics for tissue context