Peptide Mass Fingerprinting (PMF), also known as protein fingerprinting, is a high-throughput protein analysis method developed in 1933 for protein identification. An endoprotease first cleaves the unknown target protein into smaller peptide fragments, whose absolute masses can be accurately measured using a mass spectrometer, and a list of peptide peaks of the unknown protein can be obtained. Bioinformatics methods are used to compare the peak list with theoretical peptide peak lists in a protein database. The theoretical database translates all known genomes of all organisms into proteins, theoretically hydrolyzes proteins into small peptides, and calculates the absolute mass of peptides for each protein. Additionally, the peptide masses of the cleaved unknown protein are compared with the theoretical peptide masses of each protein encoded in the genome, and statistical analysis of the results can identify the best match.
Peptide Mass Fingerprinting Analysis Process
Peptide Mass Fingerprinting Analysis Process
After purification of proteins from biological tissues or cells, they are separated using 2D or SDS-PAGE gels and compared using imaging techniques. Proteins with significant differences are selected for further analysis. The selected proteins are subjected to de-gelation and enzymatic digestion to obtain a series of peptide fragments that reflect the characteristics of the protein. The peptide fingerprinting obtained from precise mass spectrometry measurements of the peptide mixture provides the molecular weight of the protein, offering accuracy for searching the selected proteins in known databases.
Applications of Peptide Mass Fingerprinting
With the advancement of analytical chemistry technologies, PMF has been widely applied in various fields such as authenticity identification of drugs, food quality control, and human disease diagnosis due to its high sensitivity, good stability, and minimal sample loss. The World Health Organization and the China National Medical Products Administration consider PMF as a technique for substance identification and process control, providing objective indicators for quality evaluation.
Advantages of Peptide Mass Fingerprinting
1. Protein identification does not rely on protein sequencing but rather on peptide mass;
2. The process is simple and quick, making it a classic method for protein identification using primary mass spectrometry.
Limitations of Peptide Mass Fingerprinting
1. The target protein sequence must exist in the target database;
2. It is suitable for single protein analysis. The presence of mixed proteins greatly increases the complexity of the analysis;
3. Peptides with similar masses greatly increase the difficulty of matching.
MALDI-TOF Peptide Mass Fingerprinting
Rapid and accurate protein identification is crucial in proteomics research. PMF can be measured using Matrix-Assisted Laser Desorption Ionization (MALDI) and Time-of-Flight (TOF) mass analyzers. It is one of the most commonly used methods in proteomics research.
MALDI uses laser irradiation to form a co-crystal film from the sample and matrix. The matrix absorbs energy from the laser and transfers it to the biomolecules. During the ionization process, protons are transferred to or acquired from the biomolecules, resulting in ionization. Therefore, MALDI is a soft ionization technique suitable for the determination of mixtures and biomacromolecules. The principle of TOF is that ions accelerate through the flight tube under the influence of an electric field, and ions are detected based on their flight time to the detector. MALDI-TOF-MS features high sensitivity, high accuracy, and high resolution, playing an important role in sample analysis in life sciences and many other fields.
Biotech company Baitaipike offers proteomics mass spectrometry identification services based on Peptide Mass Fingerprinting (PMF). Inquiries are welcome.
Bilingual Project Report in Chinese/English
In the technical report, Baitaipike will provide you with a detailed bilingual technical report in both Chinese and English, including:
1. Experimental Procedures (Chinese/English)
2. Relevant Mass Spectrometry Parameters (Chinese/English)
3. Mass Spectrometry Images
4. Raw Data
5. Analysis Results of Protein Molecular Weight and Purity
One-Stop Service for Peptide Mass Fingerprinting Analysis
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