计算溶液所需的质量、体积或浓度。
Bio-UltraBio™ 海肾萤光素酶分步检测试剂盒
有货
为什么选择此级别
BioReagent,用于化学发光 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
-20°C储存。超低温运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
概述
报告基因已被细胞生物学广泛使用以研究基因表达及其它与基因表达相关的细胞学事件，如受体活性、细胞内信号传导、mRNA 加工、蛋白质折叠及蛋白-蛋白相互作用等。海肾萤光素酶是基因表达中最常用的报告基因之一，是一种极度灵敏、响应快速且易于使用的报告基因，所催化的化学发光反应是测量基因表达最敏感的分析工具之一。
海肾萤光素酶（Renilla luciferase）是一种分子量约为36 kDa的蛋白，在氧气存在的条件下，可以催化腔肠素（Coelenterazine）氧化成Coelenteramide。在Coelenterazine氧化的过程中，会发出波长为480nm左右的生物萤光，该萤光可通过化学发光仪进行测定。
Bio-UltraBio™ 海肾萤光素酶分步检测试剂盒是一种高灵敏度、闪光型的分步法检测试剂盒。该试剂盒操作简单，检测速度快，只需把试剂盒提供的海肾萤光素酶检测底物和海肾萤光素酶检测缓冲液按照1:99的比例混合配制成海肾萤光素酶检测工作液，再取100 μL海肾萤光素酶检测工作液与20-100 μL裂解制备的细胞样品混合后即可立即进行化学发光检测，信号半衰期约5-10分钟。
注意事项：
1、 酶促反应对温度较为敏感，加样检测前需将细胞培养板和检测液平衡至室温。
2、 检测仪器选择：能够检测化学发光的仪器都适用本试剂盒的检测，但是针对相同的样品，不同检测器本底信号值和测量值均可能不同；且对于同一样本检测，不同仪器的数值不可横向比较。
3、 为防止孔间干扰，推荐使用不透明白色或黑色细胞培养板。
4、 海肾萤光素酶检测底物（100×）配制在无水乙醇中。由于无水乙醇容易挥发，有时会在初次使用时发现体积明显小于包装规格的情况，此时用无水乙醇把体积补足至包装规格，并混匀后即可使用。
5、 当酶表达量过低时，发光信号会降低，但信号稳定性会增强；当酶表达量过高时，信号半衰期会减少，但发光信号会增强。建议根据实际情况优化实验设计方案（如增加/减少质粒转染量），避免萤光素酶表达量过高/低。
使用说明：
1. 试剂准备
1） 海肾萤光素酶检测底物（100×）：每次开盖前需进行短暂低速离心
2） 检测工作液配制：根据实际使用量，以 99：1 的比例将适量的海肾萤光素酶检测缓冲液与海肾萤光素酶检测底物（100×）混匀，避光备用。
注：建议现配现用，剩余的检测试剂当次实验结束后，直接舍弃，不要留存。
2. 细胞裂解
1) 贴壁细胞：吸尽培养液。
悬浮细胞：离心收集细胞，弃上清。
2) 根据培养器皿类型，参考下表加入适量1×Lysis Buffer (用无菌水按 4:1 稀释 A 组分)：
器皿类型 | 96孔板 | 48孔板 | 24孔板 | 12孔板 | 6孔板 |
裂解液体积 (μl/孔) | 100 | 150 | 200 | 300 | 500 |
*注：若萤光素酶表达水平较低，可酌情减少裂解液用量（例如，6孔板可低至100μl/孔）。
3) 室温条件下，在微型震荡器上或静置裂解5min。
3. 加样
1）96孔板样品可直接进行后续检测，其他类型样品每孔吸取20-100μl细胞裂解样品（建议用量100μl；若样本量有限可减少用量，但同批次实验应保持一致）到检测板。
2）向各孔中加入100μl检测工作液。
4. 检测
启动化学发光检测仪或具备化学发光功能的多功能酶标仪，根据仪器说明书及实验需求进行参数设置，随后上机读取信号值。
Genetic reporters are used commonly in cell biology to study gene expression and other cellular events coupled to gene expression, such as receptor activity, intracellular signal transduction, mRNA processing, protein folding and protein:protein interactions. Renilla luciferase is one of the most commonly used reporter genes in gene expression. It is an extremely sensitive, fast-responding and easy-to-use reporter gene. The chemiluminescence reaction it catalyzes is one of the most sensitive analytical tools for measuring gene expression.
Renilla luciferase is a protein with a molecular weight of approximately 36 kDa. Under the presence of oxygen, renilla luciferase can catalyze the oxidation of Coelenterazine into Coelenteramide. During the oxidation process of Coelenterazine, bioluminescence with a wavelength of approximately 480 nm is emitted, and this fluorescence can be measured by a chemiluminescence analyzer.
Bio-UltraBio™ Two-Step Renilla Luciferase Reporter Gene Assay Kit is a highly sensitive, flash-type stepwise detection kit. The kit is simple to use and enables rapid detection. Simply mix the provided Renilla luciferase substrate with Renilla luciferase detection buffer in a 1:99 ratio to prepare the working solution. Then, combine 100 μL of this working solution with 20-100 μL of cell lysate sample, and immediately proceed with chemiluminescence detection. The signal half-life is approximately 5–10 minutes.
Precautions
1. The enzymatic reaction is temperature-sensitive. Both the cell culture plate and the detection reagent must be equilibrated to room temperature prior to assay initiation.
2. Detector Selection: Any instrument capable of measuring chemiluminescence is suitable for use with this kit. However, for identical samples, the background signal and measured values may vary between different detectors. Furthermore, readings obtained from different instruments for the same sample are not directly comparable.
3. To prevent well-to-well crosstalk, the use of opaque white or black cell culture plates is recommended.
4. Renilla luciferase substrate (100×) is prepared in anhydrous ethanol. Since anhydrous ethanol is prone to volatilization, sometimes it is observed that the volume is significantly smaller than the packaging specification during the initial use. In such cases, the volume can be replenished to the packaging specification using anhydrous ethanol, and then mixed well before use.
5. When enzyme expression is too low, the luminescent signal decreases but stability increases; when enzyme expression is too high, the signal half-life shortens but the luminescent signal intensifies. It is recommended to optimize the experimental design according to actual conditions (e.g., increasing or decreasing plasmid transfection amount) to avoid excessively high or low luciferase expression.
Instruction for use
1. Reagent preparation
1) Renilla luciferase substrate (100×): Before each opening of the container, a brief low-speed centrifugation is required.
2) Preparation of detection reagents: Based on the actual usage amount, mix an appropriate amount of Renilla luciferase detection buffer with the Renilla luciferase substrate (100×) in a 99:1 ratio. Store in the dark for later use.
Note: It is recommended to prepare the reagents immediately before use. The remaining detection reagents should be discarded at the end of the current experiment and not retained.
2. Cell lysis
1) Adherent cells: Aspirate the culture medium completely.
Suspension cells: Centrifuge to collect the cells and discard the supernatant.
2) Add an appropriate volume of 1× Lysis Buffer (prepared by diluting Component A 4:1 with sterile water) according to the culture vessel, as referenced in the table below:
Culture Vessel | 96-well plate | 48-well plate | 24-well plate | 12-well plate | 6-well plate |
Lysis Buffer Volume (μl/well) | 100 | 150 | 200 | 300 | 500 |
*Note: If luciferase expression level is low, consider using less lysis buffer (e.g., as low as 100μl/well for a 6-well plate).
3) Lyse the cells for 5 minutes at room temperature, either on a microplate shaker or by standing.
3. Sample Addition
1) The 96-well plate samples can be directly subjected to subsequent tests. For other types of samples, 20-100 μl of the cell lysate sample should be aspirated into each well (the recommended volume is 100 μl; if the sample volume is limited, the volume can be reduced, but it should be consistent within the same batch of experiments).
2) Add 100 μl of the detection working solution to each well.
4.Detection
Initiate the chemiluminescence detector or a multifunctional microplate reader equipped with chemiluminescence functionality. Set parameters according to the instrument manual and experimental requirements, then proceed to measure the signal values.
规格
别名
海肾萤光素酶报告基因检测试剂盒（闪光型） | Renilla萤光素酶报告基因检测试剂盒（闪光型） | 萤光素酶检测系统
英文别名
Renilla Luciferase Reporter Gene Assay Kit (Flash) | Renilla Luciferase Reporter Gene Assay Kit | Luciferase Assay System
规格或纯度
BioReagent, 用于化学发光
英文名称
Bio-UltraBio™ Two-Step Renilla Luciferase Reporter Gene Assay Kit
应用
Chemiluminescence
样品类型
cell or cell lysate
检测方法
Chemiluminescence
特异性
Renilla luciferase
检测仪器
酶标仪,化学发光仪
检测波长
480nm
敏感性
light-sensitive；Temperature-sensitive；Freeze-thaw sensitive
储存与运输
储存条件
-20°C储存
运输条件
超低温运输
稳定性与储存
各组分在相应的储存条件下保质期1年。
组分与储存
B1519758 | 组件 | 物理外观 | 100T | 1000T | 储存 | 单次测试使用量 |
B1519758A | 海肾萤光素酶报告基因细胞裂解液（5×） | 液体 | 10 mL | 100mL | -20℃ | 按需 |
B1519758B | 海肾萤光素酶检测缓冲液 | 液体 | 10 mL | 100mL | -20℃，避光 | 99 μL |
B1519758C | 海肾萤光素酶检测底物（100×） | 液体 | 0.1 mL | 1mL | -20℃，避光 | 1 μL |
技术文档
高级数据
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0838248 |
分析证书 | B1519758 |
技术文档和文章
溶液计算器
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