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    • 15. 发明公开
    • BESTIMMUNG DER AKTIVITÄT VON PROTEASEN
    • BESTIMMUNG DERAKTIVITÄTVON PROTEASEN
    • EP2179053A2
    • 2010-04-28
    • EP08784292.8
    • 2008-07-07
    • Papst Licensing GmbH & Co. KG
    • HOFER, Hans, WernerMEIER, Hans, Jörg
    • C12Q1/37C12M1/36
    • C12Q1/37G01N2333/96413
    • A method and apparatuses for determining the total content of proteases by means of measuring their enzyme activity after previous deinhibition is disclosed. In biological samples, lysosomal proteases are completely (e.g. in blood serum) or partly (e.g. in tissue homogenates) inhibited. A method proposed for the deinhibition entails immersing a solid support material (e.g. nylon or nitrocellulose) as plastic strip to which is linked, covalently or by adsorption, an inhibitor-binding substance which binds the inhibitor corresponding to the protease more strongly than the protease in the sample for measurement. After the deinhibition has taken place, the plastic strip is removed from the liquid sample for measurement, and the sample for measurement is passed on for measurement of the enzyme activity. Fluorogenic substrates from which the fluorogen 7-amino-4-trifluoromethylcoumarin is eliminated proved to be particularly advantageous for the activity measurement. These substrates make it possible for the sensitivity on measurement in microtitre plates with a fluorescence reader to be at least 10 times higher compared with conventional AMC substrates in blood serum, and thus for the fluorimetric determination of such enzyme activities in blood serum to be efficient with this measuring arrangement which is widely used in clinical laboratories.
    • 提供了一种测定蛋白酶活性的方法。 排除荧光素7-氨基-4-三氟甲基香豆素的荧光底物被证明对于活性测量是特别有利的。 这些底物使得可以用荧光读数器在微量滴定板中进行测量,并因此用于荧光测定血清中的这些酶活性。