Detection of Biomolecules Using PCT-DIA Proteomics

Proteomics is a pivotal technology in biological research, aimed at comprehensively analyzing the composition, structure, and dynamic alterations of the entire proteome in biological samples. In recent years, proteomics based on Data Independent Acquisition (DIA) has made significant strides, particularly when integrated with Pressure Cycling Technology (PCT), leading to notable improvements in detection sensitivity and throughput. By enhancing sample processing efficiency and improving peptide separation, PCT-DIA enables researchers to detect and quantify biomolecules with higher precision and comprehensiveness.

 

PCT-DIA proteomics combines the high-efficiency sample preparation capabilities of PCT with the unbiased data acquisition of DIA, facilitating accurate detection and quantification of proteins in complex biological samples. PCT utilizes pressure cycling to modulate the sample environment, which promotes efficient protein cleavage and peptide generation. This process significantly enhances the speed of sample processing while reducing the potential for operator-induced variability. Additionally, DIA technology, which captures all precursor ions simultaneously, circumvents the limitations of data dependency, allowing for a comprehensive dataset from a single experimental run.

 

Workflow for Biomolecule Detection Using PCT-DIA

1. Sample Preparation

Sample preparation is the foundational step in the PCT-DIA workflow and is crucial for the experiment's success. Common biological materials used include cells, tissues, or bodily fluids. To ensure protein stability and the accuracy of subsequent analyses, samples typically undergo pre-treatment, such as freezing and grinding.

 

2. Pressure Cycling Technology (PCT) Treatment

Load the extracted protein samples into PCT tubes or containers and subject them to repeated cycles of high and low pressure using the Pressure Cycling Technology (PCT) device. This process disrupts protein-protein and protein-nucleic acid interactions to enhance protein digestion and extraction. Following this, perform enzymatic digestion, such as with trypsin, under pressure cycling conditions to effectively break down the proteins into peptides.

 

3. Data Acquisition (DIA)

Separate peptides using high-performance liquid chromatography (HPLC). In DIA mode, the mass spectrometer performs full-range, unbiased data collection from all precursor ions, ensuring the detection of a comprehensive array of peptide signals.

 

4. Data Analysis

Specialized software analyzes the mass spectrometry data, allowing for the identification and quantification of proteins by constructing a protein library specific to the sample. Analyze protein abundance, identify differentially expressed proteins, and interpret biological significance.

 

Advantages of PCT-DIA Technology

1. High Sensitivity and Throughput

PCT-DIA is well-suited for processing complex biological samples, providing high peptide coverage, and enabling rapid analysis of large sample batches.

 

2. Unbiased Data Acquisition

Unlike traditional Data Dependent Acquisition (DDA), DIA captures data from all potential ions, significantly minimizing the risk of missing low-abundance proteins.

 

3. Enhanced Sample Processing Efficiency

PCT technology dramatically reduces the time required for sample processing while minimizing sample loss and contamination due to its automated nature.

 

Applications of PCT-DIA in Biomolecule Detection

PCT-DIA is widely applicable in fields such as biomedical research, drug discovery, and fundamental biological studies. Through this technology, researchers can accurately detect biomolecules across different conditions, aiding in the discovery of disease biomarkers, the identification of drug targets, and the investigation of protein interactions. The enhanced sensitivity of PCT-DIA is particularly advantageous for detecting low-abundance proteins that are difficult to identify using conventional methods.

 

PCT-DIA proteomics represents a cutting-edge approach by integrating the strengths of PCT and DIA technologies. It significantly improves the precision and efficiency of biomolecule detection, offering high sensitivity, throughput, and comprehensive data acquisition. This technology is poised to play an increasingly important role in proteomics and is expected to drive further innovations in biomolecular research.