Workflow of Olink Proteomics

Olink proteomics is a high-throughput technique leveraging protein-specific antibodies, widely used in life sciences for the simultaneous detection and quantification of numerous proteins. This method uses a unique immuno-amplification mechanism, enabling the analysis of hundreds to thousands of target proteins in a single assay, which is especially useful in clinical and basic research.

 

Olink Proteomics Workflow

1. Sample Preparation

The first step in Olink proteomics is the preparation of biological samples. Proteins are typically extracted from plasma, serum, tissues, or cell cultures. Proper collection, storage, and pre-treatment of these samples are critical to maintaining data accuracy. For plasma and serum samples, it is essential to rapidly freeze them after collection and store them at -80°C to prevent protein degradation.

 

2. Antibody Conjugation and Protein Detection

Following sample preparation, Olink's approach involves the use of two distinct antibodies that bind to specific proteins in the sample. These antibodies are each attached to unique DNA oligonucleotides. When these antibodies bind to the target proteins, they bring their attached DNA strands into close proximity. This immune-complex formation is key to achieving high specificity and sensitivity in detection.

 

3. Proximity Ligation and Amplification

When the two antibodies bind to the same protein, the DNA sequences attached to them are brought close enough for a ligation reaction to occur, forming a new DNA sequence. This process enhances detection sensitivity, ensuring that the signal is proportional to the protein concentration. This newly formed DNA is then amplified using real-time quantitative PCR (qPCR), further increasing the detectable signal and enabling precise protein quantification.

 

4. Data Collection and Analysis

The amplified DNA products are quantified via qPCR, which correlates directly with the abundance of the target proteins in the sample. One of Olink's key advantages is its multiplexing capacity, allowing simultaneous detection of up to 96 or 384 proteins in one assay. Data generated from the qPCR is processed using Olink Insight software, which provides comprehensive reports on protein concentrations and statistical analyses. This streamlined process enables researchers to efficiently interpret their results and identify significant protein expression changes.

 

5. Quality Control and Validation

Olink's workflow incorporates rigorous quality control measures at each step, including standard curves and both positive and negative controls. These ensure that the results are accurate and reproducible. After completing the assay, researchers often validate their findings by using additional methods such as ELISA or mass spectrometry to confirm key protein results.

 

The Olink proteomics workflow is designed for simplicity and efficiency, making it ideal for large-scale protein analyses. Its unique immuno-amplification technology, coupled with its advanced data analysis capabilities, offers researchers a powerful tool for high-precision proteomics. However, careful attention must be paid to sample handling and quality control to ensure the validity of the results. By mastering each step of the process, researchers can maximize the accuracy and reliability of their experiments.