RNase Residue Detection Kit

 

Product Name	Cat.No.	Spec.
RNase Residue Detection Kit	G3058-100T	100 T

 

 

Ribonuclease (RNase) is a kind of RNA hydrolase, which generally exists in the environment, and has high concentration residues in some biomaterials, enzymes and reagents, which has a very adverse effect on the related experimental operation of RNA. Therefore, it is necessary to detect the residue of RNA in any solution or biomaterial that comes into contact with RNase.

The kit contains a unique RNA modified substrate (RNase Substrate), which can quickly and sensitively detect the activity of RNase. The RNA modified substrate has a fluorescence group at one end and a quenching group at the other end. When there is no RNase, the quenching group is close to the fluorescence group, which suppresses the fluorescence of the fluorescence group to a very low level. In the presence of RNase, the RNA modified substrate is hydrolyzed, the fluorescence group and quenching group are separated in space, and the fluorescence group produces a large number of fluorescence signals. Since the fluorescence of RNA substrate increases with the increase of RNase activity, the results monitored by enzyme-labeled instrument can be Kinetically evaluated.

The kit provides RNase A as the positive control of the detection system, and can detect RNase A as low as 0.5 pg. In addition to detecting the activity of RNase A, it can also detect the activity of RNase T1, Micrococcal Nuclease, S1 Nuclease, Mung Bean Nuclease and Benzonase, etc. In addition, the kit is compatible with the DNase detection system. Because RNA substrate and DNA substrate have two different fluorescence labels, two kinds of substrates can be added to the same reaction system, and real-time analysis of RNase and DNase in a single sample can be carried out by enzyme-labeled instrument.

 

 

Shipped with wet ice and stored at -20°C. valid for up to 1 year.

 

 

Component Number	Component	G3608-50T
G3058-1	10×Reaction Buffer	2×1 mL
G3058-2	RNase A (1 U/μL)	10 μL
G3058-3	RNase Substrate	200 μL
G3058-4	ZnSO4 (100 mM)	200 μL
G3058-5	Nuclease-free Water	20 mL
Manual		One copy

 

 

1. 96-well black enzyme plate (Nuclease-free).

2. Nuclease-free pipettes and tips.

 

 

1. Before preparing reaction system:

a. Diluting a series of RNase A concentration gradients to make standard curves: RNase A (10 mg/mL) was diluted to 0.25 pg/μL, 0.5 pg/μL, 1 pg/μL and 2 pg/μL with Nuclease-free Water. Without adding RNase A as negative control, 10 μL was added to the 96-well enzyme plate, and the final amount of RNase A was 0, 2.5 pg, 5 pg, 10 pg and 20 pg.

b. Sample preparation: If the sample to be tested is liquid, it can be used directly; if the sample to be tested is solid, after sampling, it should be soaked in Nuclease-free Water to prepare liquid sample for standby.

2. Prepare the reaction system with reference to the following table:

When preparing the reaction system, please add the corresponding components of the kit to the 96-well plate according to the table below:

Component	Minus-RNase control	Plus-RNase control	Experimental samples
10×Reaction Buffer	10 μL	10 μL	10 μL
RNase A	0	10 μL	0
Sample	0	0	10 μL
RNase Substrate	2 μL	2 μL	2 μL
Nuclease-free Water	To 100 μL	To 100 μL	To 100 μL

Note: RNase Substrate should be added to the reaction system at the end, so as to avoid being degraded in advance. If the nuclease detected is Zn2+-dependent, such as Mung Bean Nuclease, S1 Nuclease, etc., add 1 μL 100 mM ZnSO4 to the reaction system.

3. Detection: The reaction system was mixed by shaking or gently blowing to ensure adequate mixing and then immediately detected using an enzyme labeler. Set the detection temperature of the enzyme-labeled instrument to 37°C (if there is no temperature setting, the enzyme activity may be low at room temperature), the excitation/emission wavelength is 492/518, the 30 min is detected continuously, and the time interval is 2 min (the detection time can be adjusted according to the RNase content, and when the content is low, the detection time can be extended to 60 min, and the interval is 5 min).

4. The amount of residual RNase A can be calculated from the standard curve.

 

 

1. Please read the Product Manual carefully before use.

2. As ribonuclease exists widely in the environment, be sure to operate it in a clean bench or biosafety cabinet to avoid contamination of samples.

3. The preparation of the reaction system needs to be operated on ice to prevent RNase Substrate from being degraded in advance.

4. To ensure the accuracy of detection, please prepare at least two duplicate holes.

5. If accurate residue detection of RNase A is not required, there is no need to make a standard curve.

6. The preparation of the reaction solution should be carried out on the ice.

7. RNase Substrate should be protected from light, and strong light should be avoided when preparing reaction solution.

8. Nuclease-free pipette tips should be used for the preparation and sub-packaging of the reaction solution to avoid cross-contamination between samples as far as possible.

 

For Research Use Only!