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Alpha Donor beads coated with nickel chelate. These beads can be used to capture His-tagged proteins and biomolecules, and can be used in conjunction with AlphaScreen, AlphaLISA, or AlphaPlex Acceptor beads to create no-wash assays for:
In a typical Alpha assay, 1 mg of Donor beads is sufficient to run 1,000-2,000 wells using a 25 µL reaction volume. Bead concentration can be adjusted for optimal performance.
|Antibody Conjugates||Nickel chelate|
|Bead Type or Core Bead Type||Alpha Donor|
|Experimental Type||In vitro|
|Product Brand Name||AlphaLISA|
|Shipping Condition||Blue Ice|
|Unit Size||5 mg|
The introduction of enzyme-linked immunosorbent assays (ELISAs) in the early 1970’s offered researchers a non-radiometric immunoassay platform without compromising sensitivity. Over the last 50 years scientists have made huge strides in disease research and drug discovery and a demand for greater assay throughput and sensitivity has evolved. In response, more robust immunoassays have been developed to address some of the limitations of the standard, colorimetric ELISA.
Find out about the most common limitations of traditional ELISAs and how different ELISA alternative technologies address these limitations.
Anti-inflammatory monoclonal antibody drugs that specifically target TNFα, such as Humira®, have been highly successful in the market. As patents expire on these top-selling drugs, effort has been placed on developing biosimilars. Biosimilars differ from small molecule generic drugs in that their chemical structure does not have to be exactly the same as the patented drug. Therefore, the FDA has stringent requirements for proving that the biosimilars have the same efficacy and safety profile as the patented drug. Companies that develop biosimilars are tasked with proving that the biosimilar shows equivalent pharmacokinetics as the patented drug.
Proving “biosimilarity” involves comparing parameters such as overall exposure, absorption, half-life, and clearance time using patient samples. Sensitive, robust, and fast assays are needed to measure these parameters. Traditional methods for detecting and quantifying these drugs in patient samples include time-consuming, wash-based ELISA and MSD methods. In contrast, AlphaLISA allows for fast, no-wash, high-throughput detection and quantification of the drug of interest in a variety of sample matrices. Here, we demonstrate the application of AlphaLISA for detecting biosimilars targeting TNFα.
Technical advancements in antibody engineering has brought about greater interest in more novel antibody therapeutic design and the emergence of new classes of antibody therapeutics called bispecific antibodies (bsAbs). The principle behind bispecific antibody design is to create an antibody / antibody fragment to two or more binding sites to help with the treatment of complex diseases.
As more bsAbs are produced as therapeutics, fast and accurate methods for functionally evaluating and characterizing the stability of these antibodies are necessary during both discovery and development stages, as well as during formulation and quality analysis.
In this application note, we demonstrate how AlphaLISA® assay technology with the EnVision® multimode plate reader can be used for bispecific antibody detection, through an example application to characterize the binding and specificity of a mouse bispecific antibody targeting mouse TIGIT and mouse PD-L1.
You will find out:
The interactions and bindingof proteins are implicated in a large number of biological processes. The needfor an efficient, highly sensitive assay to study large protein interactions is increasingly important. Alpha Technology is a highly flexible, homogeneous, no-wash assay ideal for the measurement of protein interactions and complexes as large as 200 nm in size
AlphaScreen Nickel Chelate Donor Beads at 5 mg/mL in 25 mM Hepes, pH 7.4, 100 mMNaCl supplemented with 0.05% Proclin-300 as a preservative.
Alpha has been used to study a wide variety of interactions, including protein:protein, protein:peptide, protein:DNA, protein:RNA, protein:carbohydrate, protein:small molecule, receptor:ligand, and nuclear receptor:ligand interactions. Both cell-based and biochemical interactions have been monitored, and applications such as phage display, ELISA, and EMSA (electrophoretic mobility shift assay) have been adapted to Alpha.
This guide presents the simple conversion of an ELISA or other immunoassay to an AlphaLISA® immunoassay.
AlphaScreen® and AlphaLISA® are bead-based assay technologies used to study biomolecular interactions in a microplate format. The acronym “Alpha” stands for Amplified Luminescent Proximity Homogeneous Assay. The assay does not require any washing steps. Binding of proteins or other binding partners captured on the beads leads to an energy transfer from one bead to the other, ultimately producing a luminescent signal.
Binding events between biomolecules are important components of biological processes and a number of these biomolecular interactions have been targeted for the development of novel therapeutic drugs. p53 is a transcription factor and tumor suppressor protein that is activated in response to cellular stress, and MDM2 was identified as a negative regulator that binds to p53 and tags it for ubiquitination and subsequent degradation.
The p53-MDM2 protein-protein interaction has been an excellent target for therapeutic drugs and therefore makes a good model system for developing an AlphaLISA assay to screen for inhibitors of the interaction. In this technical note, we show how to develop an assay to screen for inhibitors and how to measure a dissociation constant for moderate binding protein-protein interaction using AlphaLISA®.