Invitrogen™ Qubit™ RNA High Sensitivity (HS), Broad Range (BR), and Extended Range (XR) Assay Kits

Product Code.	Quantity	Assay	unitSize
10320093	500 assays	RNA Quantification, HS	500 assays
10174653	100 assays	RNA Quantification, BR	Each
10266793	500 assays	RNA Quantification, BR	Each
15840210	100 assays	RNA Quantification, ext range	Each
15810215	100 assays	RNA Quantification, ext range	Pack of 100
15859854	500 assays	RNA Quantification, ext range	Each
15898627	500 assays	RNA Quantification, ext range	Pack of 500
10034622	100 assays	RNA Quantification, HS	1 set

Product Code. 10320093 Supplier Invitrogen™ Supplier No. Q32855

Description

Requires

Qubit 2.0 Fluorometer and 500μL thin-walled PCR tubes

Kits enable quick and sensitive detection of low and high abundance RNA, and can distinguish between RNA and DNA, protein, and free nucleotides.

Achieve accurate and precise quantification of RNA with Qubit RNA HS, BR, and XR Assay Kits. These RNA quantification kits enable quick and selective detection of low and high abundance RNA samples, and can distinguish RNA from DNA, protein, and free nucleotides. Contaminants such as salts, solvents, or detergents are well-tolerated.

Qubit RNA HS, BR, and XR Assay Kits, designed for use with Qubit Fluorometers, are highly selective for RNA over DNA, protein, and free nucleotides. All kits provide concentrated assay reagent, dilution buffer, and pre-diluted RNA standards. Simply dilute the reagent using the buffer provided, add your sample (any volume between 1 μL and 20 μL is acceptable), and read the concentration using a Qubit Fluorometer.

Qubit RNA HS Assay Kit
The Qubit RNA HS (High Sensitivity) Assay Kit, when used with the Qubit Fluorometer, provides an accurate and selective method for the quantitation of low-abundance RNA samples.  Depending on sample volume, the assay kit is designed to be accurate for initial RNA sample concentrations of 0.2 to 200 ng/μL, providing a detection range of 4−200 ng.

Qubit RNA BR Assay Kit
The Qubit RNA BR (Broad-Range) Assay Kit, when used with the Qubit Fluorometer, provides an accurate and selective method for the quantitation of RNA samples.  Depending on sample volume, the assay kit is designed to be accurate for initial RNA sample concentrations of 0.5 to 1,200 ng/μL, providing a detection range of 10−1,200 ng.

Qubit RNA XR Assay Kit
The Qubit RNA XR (Extended Range) Assay Kit, designed for use with Qubit 4 and Qubit Flex Fluorometers, provides an accurate and selective method for the quantitation of high-abundance RNA samples. Depending on sample volume, the assay kit is designed to be accurate for initial RNA sample concentrations of 5 to 20,000 ng/μL, providing a detection range of 100−20,000 ng.

Notes
• Qubit RNA HS and BR Assay kits can be used with any Qubit Fluorometer
• Qubit RNA XR Assay Kits are designed to be used only with Qubit 4 or Qubit Flex Fluorometers
• Use with thin-wall, clear, 0.5-mL PCR tubes (Cat. No. Q32856) for the Qubit 4 Fluorometer and 8 x 200-μL tube strips (Cat. No. Q33252) for the Qubit Flex Fluorometer

Order Info

Shipping Condition: Room temperature

Specifications

• Assay: RNA Quantification, HS
• Detection Method: Fluorescence
• Excitation/Emission: 644/673
• For Use With (Equipment): Qubit Fluorometer
• No. of Reactions: 500 Reactions
• Quantitation Range: 4 to 200 ng
• Product Line: Quant-iT, Qubit
• Quantity: 500 assays
• Shipping Condition: Room Temperature

Frequently Asked Questions (FAQs)

I'm seeing other kit-related problems besides the "Standards incorrect" message with my Qubit assay. What do you suggest I try?

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.

The value is decreasing over time when using the Qubit Fluorometer. What could be causing this?

Please see our suggestions below:

Make sure that you take your reading only after incubating for at least 2 minutes (15 minutes for protein).

If you leave the assay tube in the Qubit Fluorometer and take multiple readings, the readings will go down as the tube heats up inside the instrument. If you want to take multiple readings, remove the tube from the instrument, place it in a tube rack, and allow it to equilibrate to room temperature for at least 30 seconds before rereading the tube.

You may read the sample up to 3 hours after mixing if it is protected from light. After this time, the reading will not be accurate.

Keep standards and sample tubes in the dark and protected from light in between readings.

What are the excitation/emission wavelengths for dyes in the Qubit Assays?

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

Can I make my own assay for the Qubit Fluorometer?

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).

I have a crude lysate. Will the Quant-iT and Qubit assays work?

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.

How does the accuracy and sensitivity of the Qubit quantitation assays using the Qubit fluorometer compare to a microplate reader?

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.

Can the Qubit kits give an indication of sample quality in nucleic acid samples?

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.

Can I use the Quant-iT and Qubit Kits with other fluorometers?

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.

Can I use the original Quant-iT Kits with the Qubit Fluorometer?

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.

The Qubit RNA High Sensitivity (HS), Broad Range (BR), and Extended Range (XR) Assay Kits (Cat. No. Q32855) and the Qubit dsDNA Quantification Assay Kits (Cat. No. Q32850) were both delivered at ambient temperature. Are they still usable?

Both kits are still usable if they were delivered at ambient temperature. However, once received, we recommend storing the individual components as indicated in the user guide(s).

What is the level of detection for somatic variants with the Oncomine Myeloid Research Assay GX?

Detection of somatic variants with the Oncomine Myeloid Research Assay GX has been verified down to 5% allele frequency.

Which extraction kits are compatible with Oncomine Myeloid Research Assay GX?

Here is a list of kits that are compatible with Oncomine Myeloid Research Assay GX:

Nucleic acid isolation - RNA samples
- MagMAX mirVana Total RNA Isolation Kit, Cat. No. A27828
- PureLink Total RNA Blood Kit, Cat. No. K156001
- RNaseZap RNase Decontamination Solution, Cat. No. AM9780, AM9782, AM9784

Nucleic acid isolation - DNA samples
- MagMAX DNA Multi-Sample Ultra 2.0 Kit, Cat. No. A36570
- PureLink Genomic DNA Mini Kit, Cat. No. K1820-01

What are the sample input requirements for Oncomine Myeloid Research Assay GX?

The recommended minimum sample input is 27.75 ng of DNA (1.11 ng/µL minimum concentration) and 14.25 ng of RNA (0.95 ng/µL minimum concentration).

What sample controls should I use with the Oncomine Myeloid Research Assay GX?

The recommended controls are:

DNA Controls:
- Seraseq® Myeloid Mutation DNA Mix, Cat. No. 0710-0408 (SeraCare®)
- Myeloid DNA Reference Standard, Cat. No. HD829 (Horizon™)
- AcroMetrix™ Oncology Hotspot Control, Cat. No. 969056

RNA Controls:
- Universal Human Reference RNA, Cat. No. 740000 (Agilent™)
- Seraseq® Myeloid Fusion RNA Mix, Cat. No. 0710-0407 (SeraCare®)

How many Oncomine Myeloid Research Assay GX samples can be multiplexed on a GX5 Chip?

Multiplex sequencing of up to 8 Oncomine Myeloid Research Assay GX samples (8 DNA+ 8 RNA) per lane on a GX5 Chip can be performed in a single run. There are 4 lanes on the GX5 Chip, so 32 total samples (32 DNA+ 32 RNA) can be run on one GX5 Chip (8 samples per run).

What are the components of Oncomine Myeloid Research Assay GX?

The Oncomine Myeloid Research Assay GX (Cat. No. A47857) includes the 2-pool DNA panel, the 1-pool RNA panel, Genexus Strip 1, and Genexus Strip 2-AS. The DNA panel is provided in two packs of 8 tubes (4 tubes of Myeloid DNA Pool 1 and 4 tubes of Myeloid DNA Pool 2 per pack). The RNA panel is provided in one pack of 8 tubes (Myeloid RNA Pool 1). Each primer pool in the panel is provided in pairs of tubes, where each tube pair contains one tube with primers in position 1 and one empty uncapped tube in position 2. Three 8-strip packs of the Genexus Strip 1 and Genexus Strip 2-AS (24 total of each strip) are provided with each kit.

How many samples can I run using Oncomine Myeloid Research Assay GX?

The contents of each Oncomine Myeloid Research Assay GX kit are sufficient for up to 32 samples (32 x 3‑pool reactions: 32 x 2‑pool DNA reactions and 32 x 1‑pool RNA reactions).

How many primer pools are present in the Oncomine Myeloid Research Assay GX?

The Oncomine Myeloid Research Assay GX includes 3 pools of Ion AmpliSeq oligonucleotide primers (a 2-pool DNA panel and a 1-pool RNA panel).

What is the Oncomine Myeloid Research Assay GX gene content?

The Oncomine Myeloid Research Assay GX targets key genes and fusions associated with major myeloid disorders, including acute myeloid leukemia (AML), myeloid dysplastic syndrome (MDS), myeloproliferative neoplasms (MPN), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), and juvenile myelomonocytic leukemia (JMML). The DNA panel comprises 40 key genes while the RNA panel includes a broad fusion panel of 29 driver genes, enabling detection over 600 unique fusion isotypes. See flyer for gene targets (https://assets.thermofisher.com/TFS-Assets/CSD/Flyers/Oncomine_myeloid_assay_gx_flyer.pdf).

What is the Oncomine Myeloid Research Assay GX designed for ?

The Oncomine Myeloid Research Assay GX is a comprehensive targeted next-generation sequencing (NGS) assay designed for sensitive detection of myeloid disorder-associated DNA mutations and fusion transcripts in blood and bone marrow samples.