1. MCE Kits
  2. Cell Biology
  3. Cell Analysis
  4. Reporter Gene Testing
  5. Renilla-Firefly Luciferase Dual Assay Kit

Renilla-Firefly Luciferase Dual Assay Kit 

双荧光素酶试剂盒

Cat. No.: HY-K1013
Manual SDS

MCE Renilla-Firefly Luciferase Dual Assay Kit 含有高纯度的 D-荧光素、腔肠素以及比例优化的反应缓冲液,实现哺乳动物细胞双萤光素酶报告基因检测。

Renilla-Firefly Luciferase Dual Assay Kit

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2.  同一机构(单位)同一产品试用装仅限申领一次,同一机构(单位)一年内

     可免费申领三个不同产品的试用装。

3.  试用装只面向终端客户

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  • Description

  • Storage

  • Protocol

  • Components

  • Documentation

Description
& Advantages

MCE Renilla-Firefly Luciferase Dual Assay Kit contains high-purity D-Luciferin, coelenterazine and proprietary reaction buffer. The Firefly Luciferase Working Solution is first added to the sample. This reagent contains D-Luciferin which can produce Firefly luciferase bioluminescence. Next, the Renilla Luciferase Working Solution is added to the same sample. It quenches the Firefly luciferase bioluminescence and produce Renilla luciferase bioluminescence. The light production of both reactions can be conveniently measured on a luminometer.

Publications
Storage

Stored at -20℃ protecting from light, and is stable for up to 12 months.

Protocol

1.    Material Preparation

1.1    Preparation of Lysis Buffer (1×)

Dilute Lysis Buffer (5×) 1:5 with dH2O (e.g. 1 mL Lysis Buffer + 4 mL dH2O)

1.2    Preparation of Firefly Luciferase Working Solution

Briefly centrifuge tube of Firefly Luciferase Substrate. Prepare the Firefly Luciferase Working Solution by resuspending the lyophilized Firefly Luciferase Substrate in 10 mL Firefly Luciferase Buffer.

1.3    Preparation of Renilla Luciferase Working Solution

Briefly centrifuge tube of Renilla Luciferase Substrate (125×). Prepare the Renilla Luciferase Working Solution by adding 80 μL Renilla Luciferase Substrate (125×) to 10 mL Renilla Luciferase Buffer. Mix well by inverting the tube several times.

Note: Firefly Luciferase Working Solution and Renilla Luciferase Working Solution can be dispensed into aliquots. When protecting from light and avoiding repetitive freeze-thaw cycles, the reagent is stable for one month at -20°C or for one year when stored at -70°C.

2.    Cell Lysis

2.1    Determine transfection parameters (plated cell density and subsequent incubation time) to ensure that cells are no more than 95% confluent at the desired time of lysate preparation.

2.2   Carefully remove culture medium from cells. Wash cells with wash solution, such as PBS, normal saline orserum-free culture medium, to remove residual medium.

2.3    Add proper volume of Lysis Buffer (1×). The recommended volumes of Lysis Buffer (1×) to add per well are as follows:

Multi-well Plate Lysis Buffer/Well
6-well 500 μL
12-well 200 μL
24-well 100 μL
48-well 50 μL
96-well 30 μL

Note: The volume of added Lysis Buffer (1×) can be adjusted proportionally.

2.4    Add proper volume of Lysis Buffer (1×). The recommended volumes of Lysis Buffer (1×) to add per well are as follows:

2.5    Place the culture plates on a shaker with gentle shaking at room temperature for 15 minutes.

2.6    After lysis, centrifuge at 12,000 rpm for 10 minutes at 4 °C. Transfer the supernatants to a new tube for further analysis.

3.    Luciferase Dual Assay

3.1    Add 20μL of cell lysate to a black flat-bottomed 96-well plate.

3.2    Add 100 μL of Firefly Luciferase Working Solution into each well containing cell lysate.

3.3    Immediately after adding the reagent, read the sample to capture the Firefly luciferase signals.

3.4    Add 100 μL of Renilla Luciferase Working Solution into each well from step 3.3, then immediately read the sample again to capture the Renilla luciferase signals.

3.5    Calculate the ratio of luminescence from the Firefly luciferase to the Renilla luciferase.

Note: 1) Assure Firefly Luciferase Working Solution and Renilla Luciferase Working Solution are equilibrated to room temperature before use.

          2) The luminescence values of Firefly and Renilla Luciferase should be subtracted from the corresponding background values.

Components
Contents HY-K1013-100T
Firefly Luciferase Buffer 10 mL
Firefly Luciferase Substrate (Lyophilized) 1 vial
Renilla Luciferase Buffer 10 mL
Renilla Luciferase Substrate (125×) 80 μL
Lysis Buffer (5×) 10 mL
Documentation
Help & FAQs
  • Do most proteins show cross-species activity?

    Species cross-reactivity must be investigated individually for each product. Many human cytokines will produce a nice response in mouse cell lines, and many mouse proteins will show activity on human cells. Other proteins may have a lower specific activity when used in the opposite species.

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产品名称:
双荧光素酶试剂盒
目录号:
HY-K1013
需求量: