High pH Reversed-Phase Peptide Fractionation Service
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Raw data files from LC-MS platforms
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Processed data tables (CSV or Excel format) containing peptide fractionation information
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Summary reports in PDF format, including chromatographic parameters, quality control data, and key results
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High-resolution image files (TIFF or PNG format) showing chromatograms and peak distributions
MtoZ Biolabs provides a high pH reversed-phase peptide fractionation service that integrates advanced high-pH reversed-phase chromatographic systems with high-resolution mass spectrometry platforms to achieve precise peptide separation and enrichment. This service enables efficient fractionation of complex peptide mixtures under alkaline conditions, improving protein identification depth, quantification accuracy, and detection coverage. It supports large-scale proteomic profiling, post-translational modification analysis, and biomarker discovery for both research and biopharmaceutical applications.
The Principle of High pH Reversed-Phase Peptide Fractionation
High pH reversed-phase chromatography is a separation method based on differences in peptide hydrophobicity and is complementary to conventional low-pH reversed-phase chromatography.
Under high-pH buffer conditions (typically pH 9–10), the charge state of peptides changes, and hydrophobic interactions together with ionization effects influence peptide retention behavior.
By adjusting the organic solvent gradient, more polar peptides elute first, while more hydrophobic peptides elute later.
This method divides complex peptide mixtures into multiple fractions, providing evenly distributed and orthogonal peptide components for subsequent low-pH RP-LC-MS/MS analysis.
High pH RP fractionation offers high resolution, excellent reproducibility, and strong MS compatibility. It is one of the most commonly used fractionation strategies in modern proteomics research.
High pH Reversed-Phase Peptide Fractionation Service at MtoZ Biolabs
MtoZ Biolabs, equipped with advanced high-pH reversed-phase separation systems and high-resolution mass spectrometry platforms, provides standardized peptide fractionation services to ensure experimental accuracy and reproducibility.
☑️ High-resolution fractionation system: Uses high-performance C18 columns and precise gradient control to achieve excellent peptide separation.
☑️ Orthogonal separation mechanism: Complementary to low-pH RP-LC, improving overall protein coverage and identification depth.
☑️ Multi-mode combined fractionation: Can be integrated with SCX or HILIC to build multidimensional fractionation systems.
☑️ Compatible with multiple sample types: Suitable for cell lysates, tissue extracts, plasma samples, and purified proteins.
☑️ Seamless MS connection: Fractionated samples can be directly used for LC-MS/MS analysis to ensure detection sensitivity and data quality.
Workflow of High pH Reversed-Phase Peptide Fractionation Service
1. Sample digestion: Proteins are hydrolyzed into peptides using trypsin or other specific enzymes.
2. Sample loading: Peptides are dissolved in high-pH buffer and loaded onto the C18 column.
3. Gradient elution: Peptides are separated according to hydrophobicity by increasing acetonitrile concentration.
4. Fraction collection: Eluted peaks are collected by time intervals or gradient changes.
5. Desalting and concentration: Fractions are desalted and concentrated for subsequent MS analysis.
Why Choose MtoZ Biolabs
1️⃣ Experienced technical team: Possesses extensive experience in peptide fractionation and MS sample preparation.
2️⃣ High-efficiency separation platform: Utilizes precision liquid chromatography systems to ensure reproducibility and resolution.
3️⃣ Strong orthogonality: Highly complementary to low-pH RP separation, significantly improving protein identification depth.
4️⃣ Customizable options: Offers single-dimensional or multidimensional fractionation modes (High pH RP–Low pH RP, High pH RP–SCX).
5️⃣ High compatibility: Fractionated samples are directly compatible with high-resolution MS systems such as Orbitrap, Q Exactive, and timsTOF.
Applications of High pH Reversed-Phase Peptide Fractionation Service
🔹 Comprehensive proteomics research: Increases the number and coverage of identified proteins in complex samples.
🔹 Quantitative proteomics: Optimizes fractionation uniformity in TMT, iTRAQ, and label-free experiments.
🔹 Post-translational modification studies: Provides pretreatment support for analyses such as phosphorylation, acetylation, and ubiquitination.
🔹 Biomarker discovery: Enhances the detection sensitivity of low-abundance peptides and improves data reliability.
Deliverables
1. Experimental report : Includes chromatographic parameters, fractionation profiles, and elution conditions.
2. Fractionation result tables: Record peptide distribution and elution time for each fraction.
3. High-resolution chromatograms: For visualization and data archiving.
4. Optional MS analysis services: Clients may select subsequent LC-MS/MS protein identification and quantification services.
FAQs
Q1: What is the service's general workflow?
Q2: What data formats are provided?
MtoZ Biolabs provides multiple standard data formats to ensure compatibility with various analysis and visualization tools. Typical deliverables include:
Additional data formats or customized data structures can be provided upon request to meet specific research or publication requirements.
Q3: How should I prepare my samples?
To ensure the accuracy and reproducibility of High pH Reversed-Phase Peptide Fractionation results, MtoZ Biolabs recommends the following preparation guidelines:
1. Sample type: Cell lysates, tissue extracts, plasma, and purified protein samples are acceptable.
2. Sample purity: Samples should be free of particulates, low in salt, and without detergents or organic solvents that may interfere with chromatographic separation or MS ionization.
3. Sample amount: At least 500 µL of liquid sample or 100 mg of solid material. Reserve additional material if replicate analysis is required.
4. Storage and shipping: Store biological samples at −80°C and ship on dry ice to maintain integrity.
5. Documentation: Provide detailed information such as sample origin, preparation method, buffer composition, and analysis requirements.
For more information, please refer to Sample Submission Guidelines for Proteomics and Sample Submission Guidelines for Metabolomics
If you are unsure about your sample preparation, our technical team can provide personalized pre-analysis consultation and a sample submission guide to help you optimize preparation and shipping conditions.
Start Your Project with MtoZ Biolabs
Contact us for more information, customized solutions, or quotations. MtoZ Biolabs will provide professional support and personalized analytical recommendations.
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