MRPS23 Antibody

What is MRPS23 and what is its biological function?

MRPS23 is a nuclear gene encoding a mitochondrial ribosomal protein that forms part of the small (28S) subunit of mitochondrial ribosomes. This protein plays an essential role in the translation of mitochondrial genes necessary for energy production through oxidative phosphorylation. MRPS23 is critical for proper mitochondrial function, with variants in this gene linked to mitochondrial disorders characterized by respiratory chain complex deficiencies . Patients with specific MRPS23 variants (such as c.119C>T; p.P40L) present with clinical manifestations including altered consciousness, recurrent vomiting, developmental delays, hearing impairment, hypoglycemia, lactic acidosis, and liver dysfunction, establishing MRPS23-associated disorders as inherited in an autosomal recessive pattern .

What research applications are suitable for MRPS23 antibodies?

MRPS23 antibodies are versatile tools applicable across multiple experimental techniques. They are widely used in Western Blotting (WB) for protein detection and quantification, enabling researchers to assess MRPS23 expression levels in various tissues and cell types . For quantitative analysis, these antibodies are employed in Enzyme-Linked Immunosorbent Assays (ELISA), allowing precise measurement of MRPS23 concentrations in serum, plasma, cell culture supernatants, and other biological fluids . Additionally, MRPS23 antibodies facilitate protein localization studies through Immunofluorescence (IF), Immunocytochemistry (ICC), and Immunohistochemistry (IHC), revealing the predominant cytoplasmic distribution of this mitochondrial protein . They are also valuable for Immunoprecipitation (IP) experiments investigating protein-protein interactions involving MRPS23 .

What types of MRPS23 antibodies are commercially available?

Several types of MRPS23 antibodies have been developed to meet diverse research needs. These include polyclonal antibodies raised in rabbits against specific amino acid regions (e.g., AA 50-76, AA 100-150, AA 125-175) of the MRPS23 protein . Mouse-derived polyclonal antibodies are also available . These antibodies vary in their target epitopes, with some recognizing the central region of the protein while others target specific domains . Both unconjugated antibodies and those conjugated with detection molecules such as biotin or APC (Allophycocyanin) are available, offering flexibility for different experimental designs and detection systems . The range of antibody options allows researchers to select the most appropriate tool based on their specific application, target species, and detection method.

What is the subcellular localization of MRPS23?

Immunofluorescence analyses have demonstrated that MRPS23 is predominantly localized in the cytoplasm of cells . This cytoplasmic distribution is consistent with its function as a component of mitochondrial ribosomes, as mitochondria are cytoplasmic organelles. The specific localization pattern provides important context for experimental design and interpretation, particularly when studying MRPS23 in relation to mitochondrial function or dysfunction. Understanding the normal subcellular distribution of MRPS23 is essential for interpreting any alterations in its localization that might occur in disease states or under experimental conditions .

How do mutations in MRPS23 affect mitochondrial function at the molecular level?

Mutations in MRPS23, particularly the c.119C>T; p.P40L variant, impair mitochondrial function through disruption of mitochondrial protein synthesis. In vitro assays of cultured fibroblasts from patients homozygous for this variant revealed combined respiratory chain complex deficiency, specifically demonstrating reduced activities of complexes I and IV . This molecular phenotype is consistent with compromised mitochondrial translation, as several key components of these respiratory chain complexes are encoded by mitochondrial DNA and synthesized by mitochondrial ribosomes. The structural alteration in MRPS23 likely affects the assembly or function of the mitochondrial ribosome, reducing the efficiency of protein synthesis for mitochondrially-encoded components of the electron transport chain. This molecular mechanism explains the clinical manifestations of affected patients, which typically include features of energy metabolism dysfunction such as lactic acidosis and hypoglycemia .

What evidence supports MRPS23 as a potential biomarker in cancer research?

Significant evidence points to MRPS23 as a potential biomarker in cancer research, particularly for hepatocellular carcinoma (HCC). Studies have demonstrated that MRPS23 is overexpressed at both mRNA and protein levels in HCC tissues compared to matched adjacent non-tumoral tissues . Western blotting analysis of HCC samples showed MRPS23 overexpression in 62.5% of patients (5 out of 8), consistent with elevated mRNA expression patterns . This differential expression suggests a potential role for MRPS23 in hepatocarcinogenesis. The consistent overexpression pattern indicates that MRPS23 could serve as a diagnostic or prognostic biomarker for HCC. Furthermore, the altered expression of this mitochondrial ribosomal protein may reflect metabolic reprogramming in cancer cells, connecting mitochondrial function to tumor development and progression .

How do researchers validate MRPS23 antibody specificity for rigorous experimental applications?

Validating MRPS23 antibody specificity is crucial for reliable experimental results. Multiple complementary approaches are employed for comprehensive validation. Western blotting is used to confirm that the antibody detects a single band of the appropriate molecular weight for MRPS23. Peptide competition assays, where the antibody is pre-incubated with the immunizing peptide, should abolish specific binding signals. Cross-validation with multiple antibodies targeting different epitopes of MRPS23 increases confidence in specificity . For functional validation, researchers employ genetic approaches such as siRNA knockdown or CRISPR-Cas9 knockout of MRPS23, expecting corresponding reduction in antibody signal. Immunoprecipitation followed by mass spectrometry can verify that the antibody captures the intended target. For immunohistochemical applications, staining patterns should match known expression profiles and subcellular localization of MRPS23 . These rigorous validation methods ensure that experimental findings genuinely reflect MRPS23 biology rather than antibody artifacts.

What is the evolutionary conservation of MRPS23 and how does this inform antibody selection?

Although not explicitly detailed in the search results, understanding the evolutionary conservation of MRPS23 is essential for antibody selection in comparative studies. Mitochondrial ribosomal proteins generally show reasonable conservation across vertebrates, reflecting their fundamental role in mitochondrial function. This conservation influences antibody selection, as researchers must consider cross-reactivity with MRPS23 orthologs when working with different model organisms. The available MRPS23 antibodies show varying cross-reactivity profiles; some are specific to human MRPS23, while others recognize human, mouse, and rat orthologs . When selecting an antibody for studies involving multiple species, researchers should verify the recognized epitope's conservation. For highly conserved regions, antibodies may work across species, while species-specific antibodies are preferable when studying regions with greater sequence divergence. This evolutionary perspective is particularly important for comparative studies examining MRPS23 function across different model systems.

What protocol optimizations are recommended for Western blotting with MRPS23 antibodies?

Optimizing Western blotting protocols for MRPS23 detection requires careful consideration of several parameters. For sample preparation, thorough lysis using buffers containing protease inhibitors is essential to preserve protein integrity. Protein quantification and equal loading (20-50 μg total protein per lane) are critical for comparative analyses. During SDS-PAGE, 10-12% polyacrylamide gels are typically appropriate for resolving MRPS23. For the primary antibody incubation, a 1:200 to 1:1000 dilution range is recommended, with overnight incubation at 4°C to maximize specific binding . The secondary antibody should be compatible with the primary antibody host species (rabbit or mouse, depending on the specific MRPS23 antibody used) . Enhanced chemiluminescence detection systems provide adequate sensitivity for visualizing MRPS23. Importantly, appropriate positive controls (tissues known to express MRPS23, such as liver samples) and negative controls (primary antibody omission) should be included in each experiment. For quantification, normalization to housekeeping proteins like β-actin is essential for accurate comparative analyses .

What considerations are critical for ELISA-based quantification of MRPS23?

ELISA-based quantification of MRPS23 requires attention to several critical factors for reliable results. The sandwich ELISA format is preferred, utilizing antibodies recognizing different epitopes of MRPS23—one for capture and another for detection . Sample type compatibility must be verified, as the commercial MRPS23 ELISA kits are validated for use with serum, plasma, cell culture supernatants, and other biological fluids . Standard curve preparation with recombinant MRPS23 protein at known concentrations is essential for accurate quantification. The assay typically involves multiple steps: coating microplates with capture antibody, adding samples, incubating with biotin-conjugated detection antibody, adding streptavidin-HRP, and finally developing with substrate solution . The total working time ranges from 3-5 hours, requiring careful timing. To ensure reproducibility, technical replicates are recommended, and components from different kit lots should never be mixed . Sensitivity and detection limits should be determined for each assay, with appropriate dilution series to ensure measurements fall within the linear range of the assay.

What protocol provides optimal results for immunohistochemical detection of MRPS23 in tissue sections?

For optimal immunohistochemical detection of MRPS23 in tissue sections, a carefully standardized protocol is recommended. Tissue processing should include proper fixation with 10% neutral buffered formalin followed by paraffin embedding. After sectioning (4-5 μm thickness), deparaffinization, and rehydration, antigen retrieval is critical—heating sections in sodium citrate buffer (pH 6.0) for 15 minutes enhances MRPS23 epitope accessibility . Endogenous peroxidase activity should be quenched using hydrogen peroxide solution (3%) for 10 minutes. For blocking non-specific binding, goat serum application for 30 minutes is effective . The primary MRPS23 antibody should be applied at a 1:200 dilution and incubated overnight at 4°C to maximize specific binding . After washing, biotinylated secondary antibody application for 1 hour at room temperature is followed by streptavidin-HRP incubation . Visualization with diaminobenzidine (DAB) produces a brown reaction product, with hematoxylin counterstaining highlighting tissue architecture . Appropriate positive controls (liver tissue) and negative controls (primary antibody omission) should be included in each staining run to validate results.

How should researchers design experiments to accurately quantify MRPS23 expression changes in disease models?

Designing experiments to accurately quantify MRPS23 expression changes in disease models requires a comprehensive, multi-level approach. At the mRNA level, RT-qPCR using validated MRPS23-specific primers (forward: 5′-GGTTTGACGTATATGACGCCTT-3′; reverse: 5′-CTCTAATCCGATCCTCGTGGTA-3′) provides sensitive quantification . The 2^-ΔΔCt method with appropriate reference genes (such as β-actin) enables reliable relative quantification . At the protein level, Western blotting and ELISA provide complementary data on MRPS23 expression. For tissue-level analysis, immunohistochemistry with standardized scoring systems allows assessment of expression patterns and cellular localization . Experimental designs should include appropriate controls: healthy vs. diseased samples, treated vs. untreated conditions, and time-course analyses when relevant. Statistical power calculations should determine sample sizes needed to detect biologically meaningful changes. For animal models, age, sex, and genetic background matching is essential. In cell culture models, consistent passage numbers and culture conditions minimize variability. This multi-faceted approach provides robust data on MRPS23 expression alterations in disease contexts.

How should researchers interpret discrepancies between MRPS23 mRNA and protein levels in experimental results?

Discrepancies between MRPS23 mRNA and protein levels require careful interpretation considering multiple regulatory layers. Post-transcriptional regulation through microRNAs or RNA-binding proteins may affect MRPS23 mRNA stability or translation efficiency. Post-translational modifications and protein degradation pathways can influence MRPS23 protein stability independently of mRNA levels. Technical factors must also be considered—RT-qPCR for mRNA quantification typically offers greater sensitivity than Western blotting for protein detection, potentially creating apparent discrepancies . Sample heterogeneity may contribute, as different cell populations within a tissue might express MRPS23 at varying levels. Temporal dynamics are important; protein levels may lag behind mRNA changes, so time-course experiments are valuable for correlating expression patterns. When evaluating such discrepancies, researchers should integrate multiple measurement techniques and biological replicates. In hepatocellular carcinoma studies, MRPS23 showed consistent overexpression at both mRNA and protein levels, suggesting primary regulation at the transcriptional level rather than post-transcriptional mechanisms .

What statistical approaches are most appropriate for analyzing MRPS23 expression data in disease versus control comparisons?

Statistical analysis of MRPS23 expression data requires approaches tailored to the experimental design and data characteristics. For paired samples (such as tumor and adjacent normal tissues), paired t-tests (for normally distributed data) or Wilcoxon signed-rank tests (for non-parametric data) are appropriate to detect significant differences . For independent groups, independent t-tests or Mann-Whitney U tests should be applied based on data distribution. When comparing multiple groups (such as different disease stages), ANOVA or Kruskal-Wallis tests followed by appropriate post-hoc tests are recommended. For correlations between MRPS23 expression and continuous variables (like patient age or tumor size), Pearson's or Spearman's correlation coefficients are suitable. Multiple testing correction (such as Bonferroni or false discovery rate methods) should be applied when performing numerous comparisons to control Type I error. Power analyses should determine adequate sample sizes to detect biologically meaningful differences. In survival analyses assessing MRPS23 expression's impact on patient outcomes, Kaplan-Meier curves with log-rank tests and Cox proportional hazards models are appropriate. Statistical significance is typically reported at p < 0.05, as seen in hepatocellular carcinoma studies of MRPS23 expression .

How can researchers differentiate between specific MRPS23 signals and non-specific binding in immunoassays?

Differentiating specific MRPS23 signals from non-specific binding in immunoassays requires implementation of multiple control strategies. Positive controls using samples with confirmed MRPS23 expression establish the expected signal pattern, while negative controls (samples without MRPS23 expression) demonstrate background levels. Technical negative controls (omitting primary antibody) identify non-specific secondary antibody binding. Peptide competition assays, where the antibody is pre-incubated with excess immunizing peptide, should substantially reduce specific signals while leaving non-specific binding unchanged . Cross-validation with multiple antibodies targeting different MRPS23 epitopes increases confidence in signal specificity—consistent patterns across different antibodies strongly suggest genuine MRPS23 detection. For definitive validation, genetic approaches using siRNA knockdown or CRISPR-Cas9 knockout of MRPS23 should proportionally reduce antibody signals. In Western blotting specifically, a single band of appropriate molecular weight indicates specificity, while multiple bands suggest cross-reactivity . The high sensitivity and specificity reported for MRPS23 ELISA kits, with "no significant cross-reactivity or interference between Human MRPS23 and analogues," exemplifies the stringent validation required for reliable MRPS23 detection .

What are the implications of MRPS23 expression changes for understanding mitochondrial dysfunction in disease states?

Alterations in MRPS23 expression carry significant implications for understanding mitochondrial dysfunction in various disease states. As a component of the mitochondrial ribosome, MRPS23 directly influences the translation of mitochondrially-encoded proteins essential for oxidative phosphorylation . MRPS23 upregulation in hepatocellular carcinoma suggests potential metabolic reprogramming, as cancer cells often exhibit altered mitochondrial function to support their growth and survival . The pathogenic variant c.119C>T; p.P40L in MRPS23 results in combined respiratory chain complex deficiency (particularly complexes I and IV), demonstrating how MRPS23 dysfunction directly impairs energy production . This mechanism explains the clinical manifestations observed in affected patients, including lactic acidosis, hypoglycemia, and neurological symptoms—all hallmarks of compromised cellular energetics . MRPS23 expression changes may also reflect compensatory mechanisms attempting to restore mitochondrial function in disease states. The autosomal recessive inheritance pattern of MRPS23-associated mitochondrial disorder provides insights into genetic susceptibility to mitochondrial dysfunction . Collectively, these findings position MRPS23 as a key player in mitochondrial homeostasis, with its expression changes serving as both markers and potential mediators of mitochondrial dysfunction in disease.

What emerging technologies might enhance the study of MRPS23 in research applications?

Emerging technologies promise to revolutionize MRPS23 research across multiple dimensions. Single-cell RNA sequencing can reveal cell-type-specific MRPS23 expression patterns within heterogeneous tissues, providing unprecedented resolution of expression dynamics in normal and disease states. Mass spectrometry-based proteomics enables comprehensive analysis of MRPS23 post-translational modifications and protein-protein interactions within the mitochondrial ribosome complex. CRISPR-Cas9 genome editing facilitates precise manipulation of MRPS23, allowing detailed structure-function studies to identify critical domains. Super-resolution microscopy techniques such as STORM or PALM can visualize MRPS23 localization within mitochondria at nanometer resolution, providing insights into its spatial organization. Proximity labeling methods like BioID or APEX can map the MRPS23 protein interaction network in living cells. Cryo-electron microscopy advances permit structural determination of MRPS23 within the intact mitochondrial ribosome, revealing mechanistic details of its function. These technological developments will drive deeper understanding of MRPS23 biology and potentially uncover novel therapeutic targets for mitochondrial disorders and cancers where MRPS23 expression is altered.

What research gaps remain in understanding MRPS23 function and its role in disease pathogenesis?

Despite progress in MRPS23 research, significant knowledge gaps remain. The precise molecular function of MRPS23 within the mitochondrial ribosome structure needs further elucidation—particularly how specific domains contribute to ribosome assembly, stability, and translational fidelity. The regulatory mechanisms controlling MRPS23 expression under normal and stress conditions remain poorly characterized. While MRPS23 mutations cause mitochondrial disorders , the genotype-phenotype correlation is incompletely understood, with limited information on how different mutations affect protein function and clinical manifestations. The functional consequences of MRPS23 overexpression in cancer require mechanistic investigation—whether it directly promotes tumor growth or represents a compensatory response. The potential of MRPS23 as a therapeutic target for mitochondrial disorders or cancers remains unexplored. Additionally, the evolutionary conservation of MRPS23 function across species needs systematic characterization to understand its fundamental biological roles. Addressing these research gaps will advance understanding of mitochondrial translation regulation and its implications for human disease, potentially leading to novel diagnostic and therapeutic approaches.