Antigen Information
- P01100
- 2353
- FOS
- G0S7
- Human
Assay Format
- IGF Signaling
- JAK/STAT Signaling
- MAPK Signaling
- PI3K-AKT Signaling
- PKC Signaling
- TGF-beta Signaling
- Human
- Cell Lysates
- Nuclear Extracts
- Sandwich-based
- Semi-Quantitative
Product Specifications
Introduction
Product Features
- Specific transcription factor-DNA binding assay
- Perfect alternative to EMSA
- Easy to perform in an ELISA format
- Non-radioactive assay
- High throughput (96-well plate format)
- Assay can be completed within 5 hours
Application Notes
- 96-well Strip Microplate pre-coated with DNA probes
- DNA Binding Buffer
- Positive Control Sample
- Specific Competitor DNA probe
- Non-specific Competitor DNA probe
- Assay Reagent
- DTT
- Wash Buffer
- Primary Antibody
- HRP-conjugated Secondary Antibody
- Antibody Diluent Buffer
- TMB One-Step Substrate Reagent
- Stop Solution
- Distilled or deionized water
- 100 ml and 1 liter graduated cylinders
- Tubes to prepare sample dilutions
- Absorbent paper
- Precision pipettes to deliver 2 µl to 1 ml volumes
- Adjustable 1-25 ml pipettes for reagent preparation < li="">
- Microplate reader capable of measuring absorbance at 450 nm
- Prepare all reagents and samples as instructed in the manual.
- Add 100 µl of sample or positive control to each well.
- Incubate 2 h at RT or O/N at 4 °C.
- Add 100 µl of prepared primary antibody to each well.
- Incubate 1 h at RT.
- Add 100 µl of prepared HRP-secondary antibody to each well.
- Incubate 1 h at RT.
- Add 100 µl of TMB One-Step Substrate Reagent to each well.
- Incubate 30 min at RT.
- Add 50 µl of Stop Solution to each well.
- Read at 450 nm immediately.
Typical Data
Figure 1Transcription factor assay of c-FOS from nuclear extracts of K562 cells or K562 cells treated with PMA (50 ng/ml) for 3 hr. A. Western-blot result of c-FOS from cytoplasmic and nuclear fractions. B. Transcription factor assay of c-FOS from nuclear fractions with the RayBio® Activity Assay Kit.

Figure 2Transcription factor assay of c-FOS from nuclear extracts of K562 cells or K562 cells treated with PMA (50 ng/ml) for 3 hr with the specific competitor or non-specific competitor. The result shows specific binding of c-FOS to the conserved binding site detected by using the RayBio® c-FOS TF Activity Assay Kit.

Storage/Stability
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2. 检查流程:
(1) 检查时先让患者口服一粒碳14尿素胶囊。
(2) 静坐25分钟后,受试者直接向集气瓶内呼气,患者呼气后集气瓶中的液体由粉红色变成无色为止,或者患者持续呼气时间已经达到3分钟后即可停止呼气。
(3) 向集气瓶中加入4.5ml稀释闪烁液后,加盖旋紧,置于液闪仪中进行检测。
(4) 检查过程中患者应当保持安静,剧烈运动后血中的酸碱度变化可能影响同位素标记CO2的呼出,另外在患者呼气时应当嘱咐患者注意不要将集气瓶中的液体误吸入口腔。向左转|向右转
碳14试剂检测方法
(1) 静坐25分钟后,受试者直接向集气瓶内呼气,患者呼气后集气瓶中的液体由粉红色变成无色为止,或者患者持续呼气时间已经达到3分钟后即可停止呼气。
(2) 向集气瓶中加入4.5ml稀释闪烁液后,加盖旋紧,置于液闪仪中进行检测。
(3) 检查过程中患者应当保持安静,剧烈运动后血中的酸碱度变化可能影响同位素标记CO2的呼出,另外在患者呼气时应当嘱咐患者注意不要将集气瓶中的液体误吸入口腔。
安全性
碳14尿素呼气试验应用于临床十几年,未见到明显的不良反应的报道。专业性评估报告证实碳14呼气试验对患者和操作人员的辐射危险可忽略不计,临床上可以安全使用。
mye-mail:pjlitao@tom.com
有几个问题想问问大家:
1.我看到相当多文献中,结束反应用的是抽滤,由于条件的限制,我们这里没有该装置,所以就取了少数文献中的方法,用12000G离心10min结束反应,然后沉淀TRIS-HCL洗涤3次。测量时在沉淀所在的EP管中加入闪烁液(该闪烁液的配方适用于样品含水量较少的情况),每管1ML。请问大家,这样结束反应行吗?沉淀是否应该烘干?
2.我用的液闪仪计数不是很稳定,如果进行校正,尤其是读数比较小的时候(如只有数百),以减少误差?
3.加入闪烁液后,是否应该避光放置一段时间后再测量?
谢谢!
SAMPOSCPMELAPSEDSAMPLETOTALELAPSED
TIME(MIN)TIME(MIN)
1**-1827543.90.5250.892
765315.21.5251.892
729279.52.5002.867
这是使用同一个底物,但所测得值,不知道为什麽CPM值一直往下滑?
十分感谢!!!



