Troubleshooting of ELISA Experiment: Analysis of Common Problems and Causes
ELK ELISA kitsare known for their comprehensive range of indicators and exceptional integrity, available in thousands of units. They exhibit high precision with both intra-assay and inter-assay coefficient of variation (CV) values below 10% and a cross-reaction rate of less than 5%.
These kits offer good recovery rates between 85% and 115%, high specificity with a cross-reaction rate of under 5%, and high sensitivity at the picogram (pg) level. Additionally, they come with a complete set of ready-to-use reagents, ensuring excellent integrity and ease of use.
After the good receiving of ELK’s kits, today we want to give an insight into the problems that can be experienced during the performance of the technique:
High Background/Non-Specific Staining
- Description of Results:
- After termination, the whole plate results show a uniform yellow or light color.
- The Standard curve is linear but the background is too high.
- Possible Reason:
- ELISA plate was not washed sufficiently.
- Recommendations and Precautions:
- Ensure that the same amount of Washing Solution is added to each microwell during the washing process.
- After washing, press the ELISA plate firmly on the absorbent paper to remove the residual buffer.
No Color Plate
- Description of Results:
- After the color development step, all wells of the ELISA plate are colorless.
- The positive control is not obvious.
- Possible Reason:
- Missing a reagent or a step.
- Recommendations and Precautions:
- Review the manual in detail and strictly follow the operating steps.
Light Color
Case #1:
- Description of Results:
- The Standard is normal, but the color of the sample is light.
- The visual result is normal, but the reading value of the microplate reader is low.
- Possible Reason:
- The sample to be tested may not contain strong positive samples, so the result may be normal.
- Recommendations and Precautions:
- In case of doubt, please test again.
Case #2:
- Description of Results:
- All wells, including Standard and Samples, are lighter in color.
- Possible Reason:
- Reagents and samples are not equilibrated before use.
- Recommendations and Precautions:
- All reagents and samples should be equilibrated at room temperature for about 30 minutes.
Case #3:
- Description of Results:
- Poor Repeatability
- Possible Reason:
- When adding liquid, too much remains on the medial wall of wells.
- Recommendations and Precautions:
- When adding liquid, the tip should try to add liquid along the bottom of the medial wall of wells without touching the bottom of the well.
Case #4:
- Description of Results:
- The color of plate is chaotic and irregular.
- Possible Reasons:
- Cross-contamination from manual plate washing.
- Cross-contamination when clapping.
- Recommendations and Precautions:
- When washing the plates by hand, the first 3 injections of the lotion should be discarded immediately. Set the soaking time for the next few times to reduce cross-contamination.
- Use a suitable absorbent paper towel when clapping the plate. Do not pat irrelevant substances into the well of the plate, and try not to pat in the same position to avoid cross-contamination.
By adhering to these troubleshooting steps, the reliability and accuracy of your ELISA experiments can be significantly improved.
For detailed protocols and further assistance, always refer to the specific manual provided with your ELISA kit. The following are Hot selling indicatorsof ELK, available at CymitQuimica:
- Mouse IL6(Interleukin 6) ELISA Kit
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- Mouse 4-Hydroxynonenal(HNE) ELISA Kit
- Human TNFa(Tumor Necrosis Factor Alpha) ELISA Kit
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- Rat LIF(Leukemia Inhibitory Factor) ELISA Kit
- Rat TNFa(Tumor Necrosis Factor Alpha) ELISA Kit
- Rat IL1b(Interleukin 1 Beta) ELISA Kit
On our CymitQuimicawebsite, you can browse the full selection of ELK Elisa Kits products. For more details, do not hesitate to contact our support team at support@cymitquimica.com. They are ready to provide you with outstanding personalized assistance.
