B-086 High Precision T Cell Testing Using the T-SPOT Technology: Improved Cell Yields From Lower Volumes of Blood When the T-SPOT Technology is Used With the T-Cell Select Reagent Kit for Cell Isolation

Abstract

Abstract Background T cells play a critical role in protecting humans from disease, and are a powerful diagnostic tool when used to measure immune responses to infection. However, the use of T cells in a clinical setting has been limited due to challenges associated with T cell testing methodologies. One of the key challenges has been the ability to scale up T cell testing to meet the needs of clinical laboratories. The T-SPOT technology is most widely known for the T-SPOT.TB test, an Interferon-Gamma Release Assay (IGRA) diagnostic test based on the detection of Mycobacterium tuberculosis-specific effector T cells using a single cell resolution platform (ELISPOT). The conventional method of running the T-SPOT.TB test was to use density gradient isolation on 6 mL of whole blood to isolate peripheral blood mononuclear cells (PBMCs) for high precision T cell testing. However, this testing was not accessible to some clinical laboratories due to the hands-on time associated with isolating PBMCs from whole-blood. The T-Cell Select reagent kit was developed to positively select mononuclear cells and enhance cell separation efficiency for the T-SPOT technology. The T-Cell Select reagent kit offers the opportunity to automate the T-SPOT technology which improves scalability and reliability, but it also means that an improved cell yield can be achieved from a lower volume of blood. This makes high precision T cell testing more accessible, whilst maintaining the high sensitivity and specificity of the T-SPOT technology. Methods PBMCs from 242 samples were isolated from 6 mL of whole blood, and were processed according to the T-SPOT.TB test package insert for density gradient isolation. Matched samples were used to isolate PBMCs from 3.5 mL of whole blood using magnetic bead-based positive selection according to the T-Cell Select reagent kit package insert. Results Cell yields were compared using matched samples for density gradient and the T-Cell Select reagent kit isolated cells. Cell count means were 6.6 vs 6.7 × 10^6 cell/mL for the T-Cell Select reagent kit and density gradient isolation methods, respectively. Of the 242 samples tested, two samples had insufficient yields to run the T-SPOT.TB test with the density gradient method, whilst the T-Cell Select reagent kit isolated sufficient numbers from all samples. Results of the T-SPOT.TB test had an overall agreement between the two methods of 97%, negative agreement 99% and positive agreement 94%. Conclusion When used with the T-SPOT technology, the T-Cell Select reagent kit provides an improved cell isolation methodology, requiring up to 50% lower blood volumes, and reducing the hands-on time associated with T cell testing. This makes T cell testing more accessible to labs, meaning that fewer blood samples need to be redrawn due to low cell numbers, providing an advantage in the immunosuppressed.