Caspase multiplexing: simultaneous homogeneous time-resolved quenching assay (TruPoint) for caspases 1, 3, and 6 |
| |
Authors: | Karvinen Jarkko Elomaa Annika Mäkinen Maija Liisa Hakala Harri Mukkala Veli Matti Peuralahti Jari Hurskainen Pertti Hovinen Jari Hemmilä Ilkka |
| |
Institution: | PerkinElmer Life and Analytical Sciences, Wallac Oy, P.O. Box 10, FIN-20101, Turku, Finland. jarkko.karvinen@perkinelmer.com |
| |
Abstract: | Caspases are a group of cysteine proteases involved in apoptosis and inflammation. A multiparametric homogeneous assay capable of measuring activity of three different caspases in a single well of a microtiter plate is described. Different fluorescent europium, samarium, terbium, and dysprosium chelates were coupled to a caspase substrate peptide, their luminescence properties, were analyzed, and their function in a time-resolved fluorescence quenching-based caspase 3 assay was studied. Substrates for caspases 1, 2, 3, 6, and 8 and granzyme B were also synthesized and their specificities for different caspases were determined. By selecting suitable lanthanide chelates and substrates we developed a multiparametric homogeneous time-resolved fluorescence quenching-based assay for caspases 1, 3, and 6. The assay was capable of measuring the activity of both single caspases and a mixture of three caspases mixed in the same well. |
| |
Keywords: | |
本文献已被 ScienceDirect PubMed 等数据库收录! |
|