RNaseAlarm – Lateral Flow RNase Activity Assay Kit

RNaseAlarm Lateral Flow RNase Activity Assay Kit (catalog AU2042) is a patent-pending RNase activity assay that shows whether active ribonucleases are present in liquid samples or on solid surfaces. Each kit includes 50 lateral flow dipsticks (4.5 x 60 mm) and a 96-well plate. The workflow is a 15-minute reaction at 37 °C followed by a 15-minute strip run, and strips are read by eye, by photograph or in a lateral flow strip reader.

The kit is sold with a choice of streptavidin reporter particle: Streptavidin Gold, Streptavidin Quantum Dot, Streptavidin Europium or Streptavidin Phycoerythrin. The gold version gives a visual readout; the fluorescent options suit labs that already read strips on a fluorescence-capable reader.

Key Features

• Signal-loss readout: an intact test line means no RNase was detected; a weaker or missing test line indicates RNase activity.
• Broad nuclease coverage: the manual lists RNase A, RNase T1, RNase I, micrococcal nuclease, S1 nuclease, mung bean nuclease, Denarase and Benzonase among the enzymes detected.
• Fast workflow: 15 minutes at 37 °C, then a 15-minute strip run.
• Adjustable sensitivity: the reaction time can be increased or decreased to reach the sensitivity you need.
• Built-in control line: a digoxigenin (DIG) label on the reporter particles produces a control line on every strip.
• Liquids and surfaces: test buffers, reagents and process samples, or check work surfaces.

What’s in the Kit

Each RNase activity assay kit includes:

• 50 lateral flow strips (4.5 x 60 mm dipsticks), stored at room temperature
• RNase Alarm Reaction Buffer, 2.5 mL, room temperature
• 2X RNase Alarm Running Buffer, 2.5 mL, room temperature
• RNase-free water, 1.5 mL, room temperature
• Biotin-RNA-FITC substrate, 250 µL, stored at -20 °C
• Biotin-DNA-DIG control, 250 µL, stored at -20 °C
• One 96-well plate

You will also need: micropipettes with disposable tips (25–1000 µL), a multichannel or stepper pipette (50–250 µL) or an electronic repeating pipette, a timer, a 37 °C incubator and, for numerical results, a lateral flow strip reader. All materials should be RNase-free; our DNase/RNase-free water is available for dilutions.

Who It’s For

• mRNA manufacturing: mRNA is extremely vulnerable to ribonuclease contamination, so routine checks of reagents, process samples and surfaces support quality monitoring.
• Molecular biology labs: confirm that water, buffers and enzyme preparations are RNase-free before RNA work.
• Reagent and enzyme QC groups: screen production lots for nuclease contamination with a low-equipment test.
• Inhibitor studies: apply an RNase inhibitor treatment to detect inhibition activity.

How to Use the RNase Activity Assay

1. Calculate the number of reactions and remove any strips you will not use from the 96-well plate frame.
2. Add 50 µL of RNase Alarm Reaction Buffer to each well.
3. Add 5 µL of Biotin-DNA-DIG, then 5 µL of Biotin-RNA-FITC, to each well.
4. Add 5 µL of sample, negative control or positive control to its designated well, and mix with a new RNase-free tip for each reaction.
5. Cover the plate with plastic wrap and incubate for 15 minutes at 37 °C.
6. Add 50 µL of 2X RNase Alarm Running Buffer and mix well.
7. Add one lateral flow strip to each well and run for 15 minutes.
8. Review results by photographing the strips or reading them in a lateral flow strip reader.

Full instructions and precautions are in the RNaseAlarm kit manual (PDF).

How It Works and Reading Results

The RNA substrate carries a biotin that binds the streptavidin reporter particles and a 3′ FITC/FAM group that is captured by the anti-FITC/FAM antibody at the test line. While the RNA is intact, it tethers particles to the test line and a line forms. When RNases are present, the substrate is cleaved, the two tags are separated, and the test line diminishes or disappears.

• Test line: compare each sample strip with the negative control; a decrease in test line intensity indicates RNase activity.
• Control line: produced by the DIG-labeled particles and confirms that the strip ran.
• Controls: run a negative and a positive control with every sample set so readings stay comparable across days, times and users. An RNase A spike can be used to generate the positive control.

Frequently Asked Questions

What does a missing test line mean?

In this RNase activity assay, loss of the test line means ribonucleases in the sample cleaved the RNA substrate. A test line similar to the negative control means no RNase activity was detected.

How sensitive is the test?

Sensitivity depends on the reaction time, which can be lengthened or shortened. In Attogene’s October 2023 technical article, a visible decrease in test line signal was seen with as little as 10 pg of RNase A.

Can I test surfaces as well as liquids?

Yes. The kit is designed for liquid samples and solid surfaces, and the technical article shows swabs taken from a surface with dried RNase A.

Which particle option should I choose?

Streptavidin Gold gives a visible line that can be read by eye or with a standard reader. The quantum dot, europium and phycoerythrin options are fluorescent reporters intended for labs with a matching fluorescence reader. The kit manual describes the gold format.

How should the kit be stored?

Store the Biotin-RNA-FITC and Biotin-DNA-DIG at -20 °C and everything else at room temperature. Open the RNA substrate in a clean area away from ribonuclease contamination.

Do you offer related nuclease tests?

Yes. The DNaseAlarm Lateral Flow DNase Activity Assay Kit detects DNase with the same signal-loss approach, and the RNase H Lateral Flow Activity Assay Kit measures RNase H on an RNA/DNA hybrid. To develop your own nucleic acid tests, see our NALF barcode test strips.

For research use only. Not for use in diagnostic procedures. Questions? Call 512-333-1330 or email sales@attogene.com.