- The new CRISPR-based tool, named SHERLOCK, can detect many diseases and infection like dengue, HPV and Zika.
- It can check the presence of target molecule(s) in a given sample, without requiring any additional expensive equipment.
A team of researchers from Harvard University and MIT have developed an inexpensive, highly sensitive CRISPR-based tool named SHERLOCK, and a simple paper strip that displays test results for a single genetic signature. It doesn’t require any expensive instrument – the results are visible to the naked eye.
The paper shows test results much similar to that of pregnancy test kits. It needs to be dipped into a sample and after some time, a dark horizontal line appears, showing whether the target atoms/molecule is present in the sample or not.
Along with adding the feature to precisely quantify the amount of target in a given sample, researchers increase the sensitive of the SHERLOCK, and enable it to identify multiple targets at once. Let’s dig deeper and find out what exactly they have done.
The New Version of SHERLOCK
In 2017, the team came up with SHERLOCK, stands for Specific High Sensitivity Reporter unLOCKing, that harnesses CRISPR systems (family of DNA sequences in bacteria) for beyond gene editing uses. Now they have made some advancements that accelerate its ability to rapidly and accurately detect genetic signature including tumor DNA and pathogens.
Previously, the team demonstrated the utility of SHERLOCK for a wide range of applications. Now they’ve used it to identify Dengue virus and synthetic Zika simultaneously, as well as to detect cell-free tumor DNA of lung cancer patients.
According to researchers, these improvements provide more diagnostic data and put them closer to a system that could be deployed in practical applications.
Paper Strip Results
As we’ve mentioned above, with SHERLOCk, one can check the presence of target in a sample, without using any equipment, moving us much closer to field-ready diagnostics.
Photo credit: Zhang lab/Broad Institute of MIT and Harvard
As you can see in the above image, the unused paper strips on the left doesn’t have any mark. The strips in the middle show a negative SHERLOCK readout, and right ones display a positive SHERLOCK readout.
Along with detecting mutations and diagnosing infections, the technology can be used for agricultural and industrial applications, where analyzing and observing steps along with supply chain could enhance safety and decrease waste.
Initially, SHERLOCK was able to detect only one nucleic acid sequence at a time, but now a single test can provide fluorescent signals for up to 4 different targets. For instance, it can verify whether a sample consists of dengue or Zika virus particles (both have similar symptoms) in a single reaction.
How Does It work?
The functioning of SHERLOCK is based on Cas13, a protein associated with CRISPR. One can program it to bind to a particular part of RNA. Any genetic sequence can act as a target of Cas13. These sequences include viral genomes, mutations causing cancer, and genes that confer antibiotic resistance in bacteria. To produce additional signals, SHERLOCK uses Cas12a and Cas13 enzymes from multiple species.
In specific scenarios, once Cas13 is located and cut its target, the enzyme passes to overdrive, aimlessly cutting neighbor RNA. In order to build SHERLOCK, the researchers harnessed this off-target activity, turning it to their advantage and configuring the system to support both RNA and DNA.
The diagnostic capabilities of SHERLOCK depend on additional synthetic RNA strands. Cas13 cuts this RNA if it hits the original target and releases the signaling molecule, indicating the absence or presence of target in the sample.
More specifically, SHERLOCK can be used for multiplexed genotyping for development of pharmacogenomic therapeutic, determining the presence of co-occurring pathogens and detecting genetically modified organism in the field. Also, the quick, isothermal readout provides an opportunity to detect where portable readers are absent, even for rare species like circulating DNA.
The new version of SHERLOCK is 100 more sensitive that that of the previous one. The is very crucial for applications where target concentration is extremely low, like detecting cell-free tumor DNA in blood samples. Of course, the next generation of SHERLOCK will be more precise.
In the future, it may be possible to create solution-based colorimetric readouts and multiplex lateral flow assays consisting of several test strips for different targets.
Before commercializing the technology, the researchers have to make sure that system is incredibly accurate. For developing countries that lack state-of-the-art instrument and training, these platforms could really make a difference.