Double cut test paper, suitable for CRISPR, AGO system
Single-plex CRISPR assay strips, compatible with AGO
See the picture below for the design
Two small molecule markers, FAM and biotin, were introduced at both ends of the probe. The anti-FAM antibody is labeled with colloidal gold, and the detection line on the NC membrane is coated with streptavidin that can bind biotin.
Adjust the probe concentration (recommended 100nM/L) to ensure that when the probe is not cut, all colloidal gold microspheres are captured at the red line in the above figure, the quality control line appears, the test line does not, and the result is negative.
Once the probe is cut, there will be colloidal gold microspheres crossing the red C line to the position of the blue line in the figure above, and will be captured by the anti-FAM antibody (goat anti-mouse), the test line will appear, and the result will be positive.
Highly recommented:
Double Cut Assay Strip, CRISPR, AGO Compatible
On the single-cut test paper, use goat anti-mouse at the T line position. Goat anti-mouse is non-specific to capture mouse monoclonal antibody, but because there is only one T line, it only needs to distinguish whether the probe has been cut, so it has no effect. If it is desired to realize a test strip to detect double cuts, such a design cannot be used.
As shown in the figure below, if the T1 line and T2 line still use goat anti-mouse, it is impossible to distinguish whether the captured microspheres are anti-FITC/FAM or anti-DIG.
In order to identify which probe was cut, we used aptamers that specifically capture anti-FITC/FAM and specifically capture anti-DIG, and performed reverse screening to ensure the specificity of the two aptamers. So that there will be no intersection during the test.
The figure below is the actual measurement picture, the probe addition amount is 100nM/L, and the test is carried out without CAS enzyme or AGO enzyme cleavage.
