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Updated: Sep 23, 2026
Protein Engineering by Yeast Surface Display
Published on: November 29, 2024
Recombinant antigen expression on yeast surface (RAYS) for the detection of serological immune responses in cancer
Axel Mischo1, Andreas Wadle, Kristin Wätzig
1I. Med. Klinik, Saarland University Medical School, Homburg/Saar, Germany.
Abstract:
The serological analysis of antigens by recombinant expression cloning (SEREX) has identified a multitude of new tumor antigens in many different tumor entities. These antigens can be grouped into different classes according to their specificities, with cancer/testis antigens appearing to be the most attractive candidates for vaccine development. The observation that CD8 and CD4 T-cell responses against cancer/testis antigens such as NY-ESO-1 correlate with the presence of specific antibodies demonstrates the importance of serological monitoring patients participating in vaccine trials. However, all serological assays available (Western blot, phage display and ELISA) are hampered by the fact that the protein cannot be analyzed in its natural conformation. We have thus developed a yeast display system where the antigen is expressed on the yeast surface (RAYS), allowing for a more natural folding of the protein. To validate this approach we displayed the A33 colorectal cancer antigen on the yeast cell surface and demonstrated specific binding by an A33 monoclonal antibody recognizing a conformation-dependent epitope on the A33 antigen. We then compared RAYS with the more commonly used ELISA and Western blot serological monitoring methods by analyzing 50 sera from cancer patients with known NY-ESO-1 antibody status and 10 sera from patients with unknown SSX2 antibody status in a blind fashion. RAYS appears at least equivalent to both ELISA and Western blotting for the monitoring of antibodies against NY-ESO-1 as regards specificity and sensitivity, while antibodies against SSX2 were detected more frequently by RAYS than by ELISA or phage display.
Insights
A novel yeast display system (RAYS) allows for more natural protein folding, improving cancer-testis antigen detection. This method is at least equivalent to traditional assays for monitoring patient antibody responses in vaccine trials.
Area of Science:
- Oncology
- Immunology
- Biotechnology
Background:
- Serological analysis of antigens by recombinant expression cloning (SEREX) has identified numerous tumor antigens, particularly cancer/testis antigens, as promising vaccine targets.
- Monitoring T-cell and antibody responses is crucial for cancer vaccine trials, but current serological assays (ELISA, Western blot, phage display) analyze proteins in non-natural conformations.
- This limitation hinders accurate assessment of patient immune responses to tumor antigens.
Purpose of the Study:
- To develop and validate a novel yeast display system (RAYS) for analyzing tumor antigens in their natural conformation.
- To compare the diagnostic performance of RAYS with established serological methods (ELISA, Western blot, phage display) for detecting cancer patient antibodies.
Main Methods:
- Developed a yeast display system (RAYS) to express antigens on the yeast cell surface, facilitating natural protein folding.
- Validated RAYS by displaying the A33 colorectal cancer antigen and confirming binding with an A33-specific monoclonal antibody recognizing a conformational epitope.
- Blindly compared RAYS with ELISA and Western blotting using sera from cancer patients with known NY-ESO-1 antibody status and unknown SSX2 antibody status.
Main Results:
- The RAYS system demonstrated specific binding with a conformation-dependent antibody, validating its ability to present antigens in a natural fold.
- RAYS showed sensitivity and specificity at least equivalent to ELISA and Western blotting for monitoring NY-ESO-1 antibodies.
- RAYS detected antibodies against SSX2 more frequently than ELISA or phage display, suggesting improved sensitivity for certain antigens.
Conclusions:
- The RAYS yeast display system offers a valuable platform for analyzing tumor antigens in their native conformation.
- RAYS is a robust and potentially more sensitive method for serological monitoring in cancer vaccine trials compared to traditional assays.
- This approach may enhance the evaluation of patient immune responses and the development of effective cancer immunotherapies.
