I had experienced the "Yellow background indicates a high reference wavelength read", while quantifying RNA samples (Trizol kit). Rather than DNA and cDNA samples were quantified well in the same instrument (Bioteck synergy H1 microplate reader - via Take3 plate). So, I re-quantified the RNA samples using a Nano drop, it gives precise measurements RNA samples.
What is the reason behind this?
Thanks in regards.
Hi Karthick,
Without knowing what systems you are using or the details of your assay, the yellow color (possibly Gold) means that one of your reagents could be contaminated. I would start from scratch with fresh reagents and see if this could solve your issue. I would need more information on the assays, times, systems used, etc.
Trizol kit method (2 days protocol) were used for the isolation of RNA from rat's liver sample. The RNA quantified using the Bioteck synergy H1 microplate reader - via Take3 plate.
The yellow coloration of RNA samples extracted from rat liver can be attributed to several factors:
- Phenol contamination: Trizol contains phenol, an organic solvent that can cause a yellow tint in samples. If the washing steps are not performed thoroughly, residual phenol can remain in the sample and alter its color.
- Protein contamination: Incomplete removal of proteins can interfere with the reagents used for RNA quantification and lead to a yellow hue.
- Oxidation of phenolic compounds: Prolonged exposure of samples to air can oxidize the phenolic compounds present in the sample, resulting in a yellow discoloration.
- Presence of natural pigments: Some tissues, like the liver, contain natural pigments that can contribute to the color change of the samples.
It is because sample absorbance is higher at the particular wavelength used for RNA concentration measurement. Try diluting the sample 1:1 with RNase-free water. You can further adjust the dilution based on the RNA concentration.
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