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Programmed death ligand 1 (PD-L1), also known as cluster of differentiation 274 (CD274) or B7 homolog1 (B7-H1) belongs to the growing B7 family of immune proteins and has been demonstrated to play a role in the regulation of immune responses and peripheral tolerance. Human PD-L1 is constitutively expressed in several organs such as heart, skeletal muscle, placenta and lung, and in lower amounts in thymus, spleen, kidney and liver. PD-L1, together with PD-L2, are two ligands for PD-1 (programmed death 1), a member of the CD28 family of immunoreceptors. By binding to PD-1 on activated T-cells and B-cells, PD-L1 may inhibit ongoing T-cell responses by inducing apoptosis and arresting cell-cycle progression. Accordingly, it leads to growth of immunogenic tumor growth by increasing apoptosis of antigen specific T cells and may contribute to immune evasion by cancers. PD-L1 thus is regarded as promising therapeutic target for human autoimmune disease and malignant cancers.
AlphaLISA technology allows the detection of molecules of interest in a no-wash, highly sensitive, quantitative assay. In an AlphaLISA assay, a biotinylated anti-analyte antibody binds to the Streptavidin-coated Donor beads while another anti-analyte antibody is conjugated to AlphaLISA Acceptor beads. In the presence of the analyte, the beads come into close proximity. The excitation of the Donor beads causes the release of singlet oxygen molecules that triggers a cascade of energy transfer in the Acceptor beads, resulting in a sharp peak of light emission at 615 nm.
|Assay Target Class||Protein|
|Experimental Type||In vitro|
|Product Brand Name||AlphaLISA|
|Shipping Condition||Blue Ice|
|Unit Size||100 assay points|
Too many candidates, too little time. The lack of robust, rapid, high-throughput assays to identify and qualify potential therapeutic targets in areas such as cancer research continues to cost valuable time. What if you could increase assay throughput without compromising sensitivity, obtain more data points from each sample and eliminate tedious wash steps? Find out how AlphaLISA® assay technology, combined with the EnVision® multimode plate reader, provides a fast, powerful, homogeneous platform for screening potential inhibitors of PD-L1 (a protein associated with breast cancer tumor cells) expression in human cells.
One approach to immunotherapy is the modulation of immune checkpoints that are critical in regulating the degree and duration of immune system responses and preventing autoimmunity.
In this application note, you will learn:
Breast cancer tumors can adapt to immune cell infiltration by responding to the increased concentration of interferon gamma (IFN-ɣ) and other cytokines secreted by subsets of T lymphocytes with the upregulation of the immune checkpoint proteins such as Programmed cell death ligand 1 (PD-L1). These checkpoint proteins allow the tumors to evade immune targeting and reduce the immune response, thus promoting tumor progression.
In this application note, you will learn:
When PD-1, which is expressed on the T cell, binds to PD-L1 expressed on the tumor cell, the T cell response is suppressed. Utilization of this pathway leads to tumor immune escape and promotes tumor cell growth. In fact, PD-L1 expression increases with tumor severity in many types of cancer. Release of a soluble form of PD-L1 (sPD-L1) into circulation is one mechanism that tumors may use to evade the immune response; however, it is unclear whether sPD-L1 can bind PD-1 and deliver an inhibitory signal. Previous studies have shown that soluble forms of PD-L1 have been detected in supernatants of cancer cell lines.
Traditional methods for assessing soluble and membrane-associated PD-L1 are wash-based ELISA assays, which typically require 5-6 hours of assay time. AlphaLISA® technology provides a rapid, no-wash bead-based alternative to traditional ELISAs. In this Application Note, we demonstrate how AlphaLISA is used to detect the presence of PD-L1.
Various cytokines are secreted during an active immune response that can have modulatory effects on target cell populations, including interferon gamma (IFN-ɣ), tumor necrosis factor alpha (TNFa) and several interleukins.
In this application note, you will learn how we investigated:
Immune checkpoints serve a critical role in the immune system to prevent autoimmunity and manage the degree and duration of an immune response. Cytotoxic T-Lymphocyte-associated protein 4 (CTLA-4 or CD152) is an inhibitory transmembrane protein involved in an immune checkpoint of significant interest for therapeutic development. When CTLA-4 is expressed and competes with CD28, the immune system response is downregulated. As a result of this immune system response balance, immune checkpoints provide an opportunity for therapeutic intervention to modulate immune system activity.
There is a high demand for new drugs to block CTLA-4 and modulate immune system activity. In this application note, we demonstrate how to screen for novel CTLA-4 blocking drugs by utilizing the AlphaLISA CTLA-4/CD80 binding assay.