Odyssey® 28KD Loading Indicator – 800 nm
Odyssey® 28 KD Loading Indicator – 700 nm
Evaluate Sample Loading Consistency and Uniformity with Odyssey Loading Indicators
Odyssey Loading Indicators (OLI) provide a simple, convenient method to evaluate the consistency of sample loading volume across gel lanes, as well as the uniformity of Western blot transfer.
It is essential to have consistent sample loading between gel lanes for comparative and quantitative Western blot analysis. By adding equal amounts of an Odyssey Loading Indicator, you can determine if loading was consistent or if a pipetting error occurred.
Odyssey Loading Indicators (OLI) 提供了一种简单方便的方法来评估凝胶通道上样品上样量的一致性，以及蛋白质印迹转移的均匀性。
对于比较和定量的蛋白质印迹分析，凝胶泳道之间必须有一致的上样量。 通过添加等量的 Odyssey 上样指示剂，您可以确定上样是否一致或是否发生移液错误。
- Evaluate Consistency. Ensure samples were equally loaded in each lane.
- Check Uniform Transfer. Confirm that transfer from gel to membrane was uniform.
- Assess Stable Expression. Help determine if a housekeeping protein is stably expressed and can be used for Western blot normalization.
Note: Accurate normalization relies on an internal loading control, an endogenous protein used as an indicator of sample concentration. Since the OLI is an exogenous protein, it functions as an external loading control, and cannot be used for normalization. However, you can use OLI to help check if your internal loading control is stably expressed.
Visit our Western blot Normalization page for more information on Western blot normalization strategies.
Assess Stable Expression of Housekeeping Proteins for Normalization
If you’re using a housekeeping protein to normalize Western blot data, you must validate that its expression is constant across all samples and is unaffected by experimental conditions. As an exogenous protein, the Odyssey Loading Indicator is not subject to up- or down-regulation as a function of experimental sample treatment. A stable signal for an OLI across several lanes indicates that any signal variation for the housekeeping protein was not caused by inconsistent sample loading.
如果您使用看家蛋白来标准化蛋白质印迹数据，您必须验证其在所有样品中的表达是恒定的，并且不受实验条件的影响。 作为外源性蛋白质，Odyssey Loading Indicator 不受实验样品处理的上调或下调影响。 跨多个泳道的 OLI 信号稳定表明管家蛋白的任何信号变化都不是由不一致的样品加载引起的。
“‘House-keeping’ proteins should not be used for normalization without evidence that experimental manipulations do not affect their expression.”
Figure 2. Assessing stable expression of tubulin and COX IV with OLI. The housekeeping protein signal intensity was plotted, along with signal intensities of OLI and Revert 700 Total Protein Stain. Both (A) tubulin and (B) COX IV signals varied in response to treatment, while Revert 700 Total Protein Stain signals and OLI signals remained constant. These quantitative data indicate the expression of both housekeeping proteins was affected by Etoposide treatment.
图 2. 使用 OLI 评估微管蛋白和 COX IV 的稳定表达。 绘制管家蛋白信号强度以及 OLI 和 Revert 700 总蛋白染色剂的信号强度。 (A) 微管蛋白和 (B) COX IV 信号均随治疗而变化，而 Revert 700 Total Protein Stain 信号和 OLI 信号保持不变。 这些定量数据表明两种看家蛋白的表达都受到依托泊苷处理的影响。