ELISA sample value is lower than the blank value of the problem solution - Database & Sql Blog Articles

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During ELISA experiments, it's common to encounter a situation where the sample value is lower than the blank value. This issue can be quite frustrating and often leads to confusion in interpreting results.

This problem is particularly prevalent when the sample's concentration is below the detection limit of the kit, especially with serum or plasma samples. In such cases, the measured signal might fall below the background level, resulting in inaccurate readings or even negative values.

The main causes of this phenomenon can be attributed to two primary factors: measurement errors and matrix effects.

1. Measurement Errors

Measurement errors are categorized into systematic, random, and gross errors. Among these, systematic errors do not affect the difference between the sample and blank values. However, random and gross errors are more likely to lead to sample values being lower than the blank value.

Random Errors: These occur due to uncontrollable factors and follow a normal distribution. A CV value of 20% implies that the random error could range up to ±60%. If the blank value has only one measurement, there's a higher chance that the sample value could fall below it due to random fluctuations.

To reduce random errors, you can:

• Repeat the blank measurements multiple times (usually 10 times) to get a more accurate mean value.
• Improve experimental techniques to minimize variability, especially when dealing with low concentrations.

Gross Errors: These are usually caused by human mistakes, such as incorrect pipetting, contamination, or improper washing. These errors can significantly elevate the blank value, making the sample appear lower in comparison.

Solution: Repeat the experiment and strictly follow standardized procedures to avoid such errors.

2. Matrix Effects

Matrix effects refer to the influence of other components in the sample besides the target analyte. These components can interfere with the assay, leading to inaccurate results. For example, differences in protein content, pH, or other factors between the sample and the standard solution can cause the sample to produce a weaker signal than expected.

ELISA kits are typically developed using simulated matrices rather than actual biological samples. This can result in discrepancies when testing real samples, especially if the matrix affects antigen-antibody binding.

How to address this:

• Use a calibration curve that closely matches the matrix of your samples.
• If many samples show values below the blank, consider optimizing the matrix or using a spiked recovery approach.

In summary, encountering an ELISA sample value lower than the blank value is a common challenge. Understanding the root causes—whether related to measurement errors or matrix interference—and applying appropriate solutions can help improve the accuracy and reliability of your results.

Shanghai Jiwei Biotechnology Co., Ltd. is a professional high-tech biotechnology company specializing in R&D and sales. We offer a wide range of products including ELISA kits, standards, cell lines, antibodies, biological reagents, media, and various imported reagents. Visit our website for more information: http://www.jiweibiotech.com. Welcome new and old customers to consult and purchase!