Search
Search
View additional product information for Qubit™ RNA High Sensitivity (HS), Broad Range (BR), and Extended Range (XR) Assay Kits - FAQs (Q10210, Q10211, Q32855, Q32852, Q33224, Q33223)
11 product FAQs found
Here are several suggestions:
1.View the raw fluorescence value (RFU) for the standards under “Check Standards” or “Check Calibration”. Confirm that the values for the samples fall between the values of the standards (or a little above the highest standard). If they do not, the sample is out of the accurate range of the assay. Refer to the confidence ranges for each assay in the product manuals. The readout in the assay will be to 2 significant figures instead of 3 if the assay sample is out of the high confidence range.
To bring the sample into the accurate range, dilute the sample or use more or less of it (for example, 10 µL instead of 2 µL if the sample reads low).
2.Check for temperature issues: The assay is temperature sensitive and the fluorescent signal can decrease at higher temperatures. Temperature fluctuations between samples, or between samples and standards, can cause problems.
Make sure that the buffer and Qubit reagent in DMSO are at room temperature. The buffer and Qubit reagent should be stored at room temperature, not in the refrigerator. Even after 2-3 hours at room temperature, buffer previously stored at 4°C can remain below room temperature.
Make sure your samples and working solution are not too warm (including those straight from a centrifuge). Samples kept in the Qubit instrument too long or read multiple times can warm up. If you want to perform multiple readings of a single tube, you should remove the tube from the instrument and let it equilibrate to room temperature for 30 seconds before taking another reading. Also, do not hold tubes in your hand for very long before reading them in the instrument, since this can warm the sample, resulting in a low reading.
3.Ensure that you have prepared the Qubit working solution correctly (1:200 dilution using the buffer provided in the kit). Ensure that you have prepared the standard tubes correctly (10 µL of each standard in 190 µL of the working solution). Ensure that the tubes are filled with at least 200 µL (both standards and samples).
4.Ensure that the reagents and standards you are using are less than 6 months old, and that the standards have been stored correctly. The Qubit reagent stock solution should be protected from light as much as possible.
5.Ensure that you have selected the correct assay on the Qubit Fluorometer for the Qubit assay you are performing.
6.Ensure that the lid is completely closed when reading standards and samples.
7.Use recommended tubes (both so the tube does not obstruct the lid, and for optical clarity). Some types of tubes can have high autofluorescence that will affect the assay.
8.Did you enter the number of microliters of stock you pipetted into the working solution into the Qubit instrument? If so, the reading after giving the Qubit Fluorometer this information is the concentration of your stock solution. If you did not, the reading you got is for the concentration in the assay tube (the tube you put into the Qubit Fluorometer) and not your stock solution.
9.If you are comparing Qubit assay results to concentration obtained by UV absorbance, and the concentration based on absorbance is significantly higher, it may be because of nucleic acid or protein contamination. The Qubit assays are much more specific for DNA, RNA, or protein than absorbance readings.
Please see our suggestions below:
The exact excitation/emission wavelength information is proprietary. Here are the approximate excitation/emission wavelengths:
- Qubit dsDNA HS Assay: ~500 nm/ ~530 nm
- Qubit dsDNA BR Assay: ~510 nm/ ~530 nm
- Qubit ssDNA Assay: ~490 nm/ ~520 nm
- Qubit RNA HS Assay: ~640 nm/ ~670 nm
- Qubit RNA BR Assay: ~640 nm/ ~670 nm
- Qubit microRNA Assay: ~500 nm/ ~520 nm
- Qubit Protein Assay: ~470 nm/ ~570 nm
Find additional tips, troubleshooting help, and resources within our Nucleic Acid Quantification Support Center.
Yes, you can, for Qubit instruments developed after the original Qubit (1.0) Fluorometer. See MyQubit assay instructions here (http://www.thermofisher.com/us/en/home/life-science/laboratory-instruments/fluorometers/qubit/qubit-assays/myqubit.html.html).
Generally, the cleaner the sample the better. Some salts, proteins, and detergents are tolerated in the assays; see the specific assay protocol for which ones and at what concentrations.
The accuracy and sensitivity of the Qubit quantitation assays are the same as that of a microplate reader. This was a requirement during product development. The detection limits for each Qubit kit can be found on the corresponding product manual, which can be found by searching our website by keyword or catalog number.
Find additional tips, troubleshooting help, and resources within our Nucleic Acid Quantification Support Center.
No. The Qubit DNA and RNA kits only quantify the amount of either DNA or RNA in the sample. The Qubit fluorometer cannot take absorbance readings to provide a A260/A280 ratio or detect protein in nucleic acid samples. This can be done with the NanoDrop instrument. If your sample contains protein or other contaminants that can affect the assay, it should be further purified.
If your sample may contain both DNA and RNA, one may use either (or both) the DNA and RNA Qubit kits and compare with samples treated with either RNase or DNase to get an accurate determination of DNA or RNA, respectively.
All Quant-iT and Qubit kits are compatible with all fluorometers and microplate readers that have the appropriate light sources and filters. You won't have access to the algorithm in the Qubit fluorometer for generating the standard curve provided by the instrument, instead, you must make a few dilutions of the highest standard DNA or RNA (Standard #2) in the Qubit kits to generate a standard curve with multiple data points.
No, we do not recommend this. Some of the dyes in the original Quant-iT kits (those NOT listed as “for use with the Qubit fluorometer”) are not compatible with the Qubit Fluorometer. In addition, the new Quant-iT kits (labeled as “for use with the Qubit Fluorometer”) have standards formulated to be compatible with the Qubit fluorometer internal algorithms for the respective assays. The Qubit Fluorometer-compatible kits are also less expensive per assay if you are processing fewer than 20 samples at a time.
When stored as directed, the Qubit RNA Quantification, broad range (BR) Assay Kit (Cat. No. Q10210) is stable for up to 6 months from the date of receipt. The storage conditions are as follows:
When stored as directed (2-8 degrees C, protected from light), the Qubit RNA BR Assay Kit is stable for six months. Please refer to the user guide (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/Qubit_RNA_BR_Assay_UG.pdf) for further technical details.