{"product_id":"revvity-alsu-tcasp3-a500","title":"Revvity, ALSU-TCASP3-A500, AlphaLISA Human Total Caspase-3 Detection Kit, 500 Assay Points","description":"\u003cp\u003eOverview\u003cbr\u003e\nCaspase-3 is a cysteine-aspartic acid protease that plays a pivotal role in the execution phase of apoptosis (programmed cell death). It is synthesized as an inactive proenzyme and becomes activated through cleavage by upstream caspases in response to apoptotic stimuli. Activated Caspase-3 orchestrates apoptosis by cleaving various cellular proteins, including PARP, ICAD, and cytoskeletal elements. While critical for normal development and immune homeostasis, aberrant Caspase-3 activity is implicated in pathologies such as neurodegeneration, ischemia, and cancer. Its selective activation or inhibition is a potential therapeutic approach for diseases involving defective apoptotic processes.\u003cbr\u003e\nThe AlphaLISA SureFire Ultra Human Total Caspase-3 Detection Kit is a sandwich immunoassay for the quantitative detection of total Caspase-3 in cellular lysates, using Alpha Technology.\u003cbr\u003e\nFormats:\u003c\/p\u003e\n\n\u003cp\u003eThe HV (high volume) kit contains reagents to run 100 wells in 96-well format, using a 60 μL reaction volume.\u003cbr\u003e\nThe 500-point kit contains enough reagents to run 500 wells in 384-well format, using a 20 μL reaction volume.\u003cbr\u003e\nThe 10,000-point kit contains enough reagents to run 10,000 wells in 384-well format, using a 20 μL reaction volume.\u003cbr\u003e\nThe 50,000-point kit contains enough reagents to run 50,000 wells in 384-well format, using a 20 μL reaction volume.\u003c\/p\u003e\n\n\u003cp\u003eAlphaLISA SureFire Ultra kits are compatible with:\u003c\/p\u003e\n\n\u003cp\u003eCell and tissue lysates\u003cbr\u003e\nAntibody modulators\u003cbr\u003e\nBiotherapeutic antibodies\u003c\/p\u003e\n\n\u003cp\u003eAlphaLISA SureFire Ultra kits can be used for:\u003c\/p\u003e\n\n\u003cp\u003eCellular kinase assays\u003cbr\u003e\nReceptor activation studies\u003cbr\u003e\nHigh-throughput screening for preclinical studies\u003cbr\u003e\nHow it works\u003cbr\u003e\nTotal-AlphaLISA SureFire Ultra assay principle\u003c\/p\u003e\n\n\n    \u003cp\u003eThe Total-AlphaLISA SureFire Ultra assay measures the expression level of a protein target in a cell lysate.\u003c\/p\u003e\n\n\n    \u003cp\u003eThe Total-AlphaLISA SureFire Ultra assay uses two antibodies which recognize two different distal epitopes on the targeted protein. AlphaLISA assays require two bead types: Acceptor and Donor beads. Acceptor beads are coated with a proprietary CaptSure™ agent to specifically immobilize the assay specific antibody, labeled with a CaptSure tag. Donor beads are coated with streptavidin to capture one of the detection antibodies, which is biotinylated. In the presence of targeted protein, the two antibodies bring the Donor and Acceptor beads in close proximity whereby the singlet oxygen transfers energy to excite the Acceptor bead, allowing the generation of a luminescent Alpha signal. The amount of light emission is directly proportional to the quantity of protein present in the sample.\u003c\/p\u003e\n\n\n\n  \u003cbr\u003e\n        \n\n\n\t\u003cbr\u003e\n  \u003cbr\u003e\n        \u003cbr\u003e\n  \n\n      \u003cbr\u003e\n  \u003cbr\u003e\n  \u003cbr\u003e\n    \u003cbr\u003e\n      \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n      \u003cbr\u003e\n    \n\n    \u003cbr\u003e\n      \n\n      \u003cbr\u003e\n        \n\n      \u003cbr\u003e\n    \n\n  \n\n\n\n\n\n    \u003cp\u003e \u003c\/p\u003e\n\n\n\n\n            \u003cp\u003eTotal-AlphaLISA SureFire Ultra two-plate assay protocol\u003c\/p\u003e\u003cbr\u003e\n        \n\n            \u003cp\u003eThe two-plate protocol involves culturing and treating the cells in a 96-well plate before lysis, then transferring lysates into a 384-well OptiPlate™ plate before the addition of Total-AlphaLISA SureFire Ultra detection reagents. This protocol permits the cells viability and confluence to be monitored. In addition, lysates from a single well can be used to measure multiple targets.\u003c\/p\u003e\u003cbr\u003e\n        \n\n\n\n\n  \u003cbr\u003e\n        \n\n\n\t\u003cbr\u003e\n  \u003cbr\u003e\n        \u003cbr\u003e\n  \n\n      \u003cbr\u003e\n  \u003cbr\u003e\n  \u003cbr\u003e\n    \u003cbr\u003e\n      \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n        \u003cbr\u003e\n      \u003cbr\u003e\n    \n\n    \u003cbr\u003e\n      \n\n      \u003cbr\u003e\n        \n\n      \u003cbr\u003e\n    \n\n  \n\n\n\n\n\n\n\n\n\n            \u003cp\u003eTotal-AlphaLISA SureFire Ultra one-plate assay protocol\u003c\/p\u003e\u003cbr\u003e\n        \n\n            \u003cp\u003eDetection of Total target protein with AlphaLISA SureFire Ultra reagents can be performed in a single plate used for culturing, treatment, and lysis. No washing steps are required. This HTS designed protocol allows for miniaturization while maintaining AlphaLISA SureFire Ultra quality.\u003cbr\u003e\nAssay versatility\u003cbr\u003e\nExpression of Caspase 3 in various cell lines\u003cbr\u003e\nAdherent cells were seeded at 40,000 cells\/well in a 96-well culture plate in complete medium and incubated overnight at 37°C, 5% CO2. Cells were lysed with 100 µL of Lysis Buffer.\u003cbr\u003e\nSuspension cells were seeded at 400,000 cells\/well in a 96-well culture plate in HBSS + 0.1% BSA, cells were spun down and lysed with 100 µL of Lysis Buffer.\u003cbr\u003e\nCaspase 3 Total levels were evaluated by AlphaLISA SureFire Ultra. For the detection step, 10 µL of cell lysate (approximately 4,000 adherent cells or 40,000 suspension cells) was transferred into a 384-well white OptiPlate, followed by 5 µL of Acceptor Mix and incubated for 1 hour at RT. Finally, 5 µL of Donor Mix was then added to each well and incubated for 1 hour at RT in the dark. The plate was read on an Envision using standard AlphaLISA settings.\u003cbr\u003e\nAs expected, high Caspase 3 expression was observed in HEK293, Hep G2 and RT4 cells while expression was lower in MCF7 cells. \u003cbr\u003e\nApplication-Cell Signaling\u003cbr\u003e\nAutomation Compatible-Yes\u003cbr\u003e\nBrand-AlphaLISA SureFire Ultra\u003cbr\u003e\nDetection Modality-Alpha\u003cbr\u003e\nProduct Group-Kit\u003cbr\u003e\nProtocol Time-2h at RT\u003cbr\u003e\nSample Volume-10 µL\u003cbr\u003e\nShipping Conditions-Shipped in Blue Ice\u003cbr\u003e\nTarget-Caspase-3\u003cbr\u003e\nTarget Class-Phosphoproteins\u003cbr\u003e\nTarget Species-Human\u003cbr\u003e\nTechnology-Alpha\u003cbr\u003e\nTherapeutic Area-Inflammation Oncology\u003cbr\u003e\nUnit Size-500 assay points\u003c\/p\u003e","brand":"Revvity","offers":[{"title":"Default Title","offer_id":46774589489321,"sku":"ALSU-TCASP3-A500","price":0.99,"currency_code":"USD","in_stock":true}],"url":"https:\/\/iright.com\/products\/revvity-alsu-tcasp3-a500","provider":"Iright","version":"1.0","type":"link"}