荧光素酶活性检测试剂盒

简介：
迈晨公司开发生产的“荧光素酶活性检测试剂盒”The Luciferase Assay Kit provides a sensitive, quick, and quantitative measurement of the activity of firefly luciferase produced in cultured mammalian cells. The kit contains a cell lysis buffer, an assay buffer, and the luciferase substrate. The cell lysis buffer allows efficient extraction of luciferase from many types of cultured cells within 5 minutes. The luciferase assay is performed by adding the luciferase substrate to the cell lysate premixed with the assay buffer. Luciferase in the cell lysate catalyzes the chemiluminescent reaction, which emits light that is readily measurable with a luminometer.

试剂盒组分：

试剂盒提供试剂        Quantity       
注： 该试剂盒提供的试剂能够满足100次分析，单次分析是指24孔或者12孔细胞培养板的一孔
细胞裂解液（Extraction Buffer）        10mL       
分析试剂（Assay Buffer）        30mL       
荧光素底物（Substrate）        10mL       

操作步骤：

一、细胞裂解
        从细胞培养板中吸去培养液
        每孔中加入100微升细胞裂解液（Extraction Buffer ）
        于室温在摇床上摇动5－10分钟
        将细胞培养板放置在冰上备用

二、荧光强度测定

1. Single-Tube Measurement with Manual Injection

        Program the luminometer to perform a 2-second measurement delay followed by a 10-second measurement read for luciferase activity. The read time may be shortened if sufficient light is produced.
        Dispense 300μl of the Luciferase Assay Buffer into luminometer tubes, one tube per sample.
        Add 90μl of cell lysate to a luminometer tube containing the Luciferase Assay Buffer. Mix by pipetting 2–3 times or vortex briefly.
        Add 100μl of Substrate to a luminometer tube containing the Luciferase Assay Buffer and cell lysate. Mix by pipetting 2–3 times or vortex briefly.
        Place the tube in the luminometer and initiate reading.
        Record the reading.

2. Single-Tube Measurement with Automatic Injection

        Program the luminometer to perform a 2-second measurement delay followed by a 10-second measurement read for luciferase activity.
        Prime the luminometer injector at least three times with Luciferase Substrate Reagent.
        Dispense 300μl of the Luciferase Assay Buffer into luminometer tubes, one tube per sample.
        Add 90μl of cell lysate to a luminometer tube containing the Luciferase Assay Buffer. Mix by pipetting 2–3 times or vortex briefly.
        Place the tube in the luminometer and initiate reading by injecting 100μl of Substrate into the tube.
        Record the reading.

1.        Multi-Tube Measurement with Automatic Injection

        Program the luminometer to perform a 2-second measurement delay followed by a 10-second measurement read for luciferase activity.
        Program the luminometer with at least three injections with Luciferase Substrate.
        Dispense 300μl of the Luciferase Assay Buffer into luminometer tubes, one tube per sample.
        Add 90μl of cell lysate to a luminometer tube containing the Luciferase Assay Buffer.
        Place the tubes in the luminometer chain and initiate reading by injecting 100μl of Substrate into the tube.
        Record the reading.


2.        Plate-Reading Luminometers with Automatic Injection

The following protocol was designed for 96-well plate format:

        Program the luminometer to perform a 2-second measurement delay followed by a 10-second measurement read for luciferase activity.
        Dispense 300μl of the Luciferase Assay Buffer into each well, one well per sample.
        Add 90μl of cell lysate to the well containing the Luciferase Assay Buffer.
        Place the plate in the luminometer and initiate reading by injecting 100μl of Substrate into the well.
        The plate is advanced to the next well for a repeat of the injection and reading.
        Record the reading.

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