MRPS27 Antibody

What is MRPS27 and what is its cellular function?

MRPS27 (mitochondrial ribosomal protein S27) is an essential component of the mitochondrial ribosome involved in protein synthesis within the mitochondria. The protein plays a crucial role in the translation of mitochondrial genes. Dysregulation of MRPS27 has been associated with mitochondrial dysfunction and may contribute to several pathological conditions including cancer, metabolic disorders, and neurodegenerative diseases . The protein is primarily localized in the mitochondrion and has a calculated molecular weight of 48 kDa, which corresponds to its observed molecular weight in experimental conditions .

What applications have MRPS27 antibodies been validated for?

MRPS27 antibodies have been extensively validated for multiple experimental applications. The polyclonal antibody (17280-1-AP) from Proteintech has been validated for Western Blot (WB), Immunohistochemistry (IHC), Immunofluorescence (IF/ICC), Immunoprecipitation (IP), and ELISA applications . The recommended dilutions vary by application:

These applications have been supported by numerous publications, with Western Blot being the most commonly used technique (26 publications cited) .

What species reactivity has been confirmed for MRPS27 antibodies?

MRPS27 antibodies show reactivity with samples from multiple species. The polyclonal antibody 17280-1-AP has been tested and confirmed to react with human, mouse, and rat samples . The monoclonal antibody 66724-2-PBS has been specifically tested for human reactivity . Positive samples for antibody testing include various cell lines (A431, HepG2, A-549, LO2, U-87MG, HeLa, HT-29) and tissues (mouse brain, mouse liver, rat heart) .

What are the optimal conditions for MRPS27 detection in immunohistochemistry applications?

For optimal MRPS27 detection in IHC applications, antigen retrieval is a critical step. Based on experimental data, the recommended approach is to use TE buffer at pH 9.0 for antigen retrieval. Alternatively, citrate buffer at pH 6.0 may also be used, though this may result in different staining patterns or intensities . The recommended antibody dilution range for IHC is 1:50-1:500, but researchers should optimize this dilution based on their specific sample type and experimental conditions. Mouse brain tissue has been positively tested for MRPS27 detection in IHC applications . For all applications, it is strongly recommended to titrate the reagent in each testing system to obtain optimal results, as outcomes can be sample-dependent .

How should researchers approach MRPS27 antibody validation in knockdown/knockout experiments?

Knockdown/knockout validation is a crucial step in confirming antibody specificity. The MRPS27 antibody (17280-1-AP) has been cited in at least one publication involving KD/KO experiments, indicating its utility in such validation approaches . When designing KD/KO experiments for MRPS27, researchers should:

• Use appropriate controls (both positive and negative)

• Consider the impact of MRPS27 depletion on mitochondrial function

• Validate the knockdown efficiency using multiple techniques (e.g., qPCR and Western blot)

• Use proper statistical analysis to quantify changes in expression levels

This approach ensures robust validation of antibody specificity while also providing insights into the biological function of MRPS27.

What are the considerations for setting up a multiplex assay using MRPS27 antibodies?

For researchers interested in multiplex assays, the monoclonal antibody 66724-2-PBS is specifically designed for such applications. This antibody is available as part of matched antibody pairs validated in Cytometric bead array applications :

This antibody is provided in PBS only (BSA and azide free) storage buffer at a concentration of 1 mg/mL, making it ready for conjugation. The conjugation-ready format makes these antibodies ideal for ELISAs, multiplex assays requiring matched pairs, mass cytometry, and multiplex imaging applications . Researchers should optimize antibody use for each specific application and assay.

What storage conditions are optimal for maintaining MRPS27 antibody activity?

Proper storage is crucial for maintaining antibody activity. For the polyclonal antibody (17280-1-AP), the recommended storage condition is -20°C, where it remains stable for one year after shipment. Aliquoting is unnecessary for -20°C storage. The antibody is supplied in PBS with 0.02% sodium azide and 50% glycerol at pH 7.3 . For smaller sizes (20μL), the antibody contains 0.1% BSA .

For the monoclonal antibody (66724-2-PBS), which is provided in PBS only without BSA and azide, storage at -80°C is recommended to maintain activity . This difference in storage conditions highlights the importance of following specific recommendations for each antibody formulation.

How should researchers address background issues in Western blots using MRPS27 antibodies?

When encountering background issues in Western blots, researchers should consider several optimization strategies:

• Dilution optimization: Test the recommended dilution range (1:500-1:2000) to find the optimal concentration for your specific sample.

• Blocking optimization: Adjust blocking conditions (buffer composition, incubation time) to reduce non-specific binding.

• Sample preparation: Ensure proper sample preparation to maintain protein integrity while minimizing contamination.

• Positive controls: Include validated positive controls such as A431 cells, which have been confirmed to express detectable levels of MRPS27 .

• Washing steps: Optimize washing steps (buffer composition, duration, number of washes) to reduce background while maintaining specific signal.

What are the critical considerations for immunofluorescence experiments targeting MRPS27?

For successful immunofluorescence experiments targeting MRPS27, researchers should consider the following:

• Cell types: HepG2 cells have been positively tested for IF/ICC applications , while other cell lines like A-549, LO2, U-87MG, HeLa, A-431, and HT-29 have been used as positive samples .

• Antibody dilution: The recommended dilution range for IF/ICC is 1:200-1:800 for the polyclonal antibody (17280-1-AP) and 1:50-1:200 for another antibody formulation (CAB11667) .

• Fixation method: Select an appropriate fixation method that preserves mitochondrial structure while maintaining antigen accessibility.

• Co-staining: Consider co-staining with mitochondrial markers to confirm the expected mitochondrial localization of MRPS27.

• Image acquisition: Use appropriate microscopy settings to capture the mitochondrial distribution pattern of MRPS27.

How can MRPS27 antibodies be utilized to study mitochondrial dysfunction in disease models?

MRPS27 antibodies can be powerful tools for investigating mitochondrial dysfunction in various disease models:

• Expression analysis: Quantify MRPS27 expression levels in diseased versus healthy tissues using Western blot or IHC to identify potential alterations associated with pathological conditions.

• Localization studies: Use immunofluorescence to examine potential changes in the subcellular distribution of MRPS27 in disease models.

• Protein interaction networks: Employ immunoprecipitation (using 0.5-4.0 μg antibody for 1.0-3.0 mg of total protein lysate ) to identify altered protein-protein interactions involving MRPS27 in disease states.

• Functional studies: Combine MRPS27 antibody-based detection with functional assays of mitochondrial activity to correlate protein levels with organelle function.

Since dysregulation of MRPS27 has been associated with mitochondrial dysfunction in cancer, metabolic disorders, and neurodegenerative diseases , these approaches can provide valuable insights into disease mechanisms and potential therapeutic targets.

What role can MRPS27 immunodetection play in cancer research?

MRPS27 immunodetection can contribute significantly to cancer research through several approaches:

• Biomarker potential: Assess MRPS27 expression in various cancer types using tissue microarrays and IHC to evaluate its potential as a diagnostic or prognostic biomarker.

• Metabolic reprogramming: Investigate the relationship between MRPS27 levels and cancer-associated metabolic changes by combining antibody-based detection with metabolic profiling.

• Therapeutic targeting: Use MRPS27 antibodies to screen for compounds that modulate its expression or function as potential therapeutic interventions.

• Cancer cell line characterization: Profile MRPS27 expression across cancer cell line panels to identify correlations with other cancer-related phenotypes.

Given that mitochondrial dysfunction is a hallmark of many cancers, and MRPS27 plays a crucial role in mitochondrial protein synthesis, these studies can provide new insights into cancer biology and potential therapeutic strategies.

How might MRPS27 antibodies be used in studies of mitochondrial ribosome assembly?

MRPS27 antibodies can be valuable tools for investigating mitochondrial ribosome assembly:

• Co-immunoprecipitation: Use the antibody to pull down MRPS27 and identify its interacting partners within the mitochondrial ribosome complex.

• Sucrose gradient analysis: Combine antibody detection with sucrose gradient fractionation to track MRPS27 distribution across different ribosomal assembly intermediates.

• Pulse-chase experiments: Employ antibody detection in pulse-chase experiments to monitor the kinetics of MRPS27 incorporation into mitochondrial ribosomes.

• Structural studies: Use immunogold labeling with MRPS27 antibodies in electron microscopy to localize the protein within the mitochondrial ribosome structure.

These approaches can provide insights into the molecular mechanisms of mitochondrial ribosome assembly and the specific role of MRPS27 in this process, contributing to our understanding of mitochondrial gene expression regulation.