Utilizing CRISPR/Cas Technology for Point-of-Care Diagnosis

CRIPSR/Cas complexes possess an unparalleled ability to find specific nucleotide sequences in vast amounts of genomic material. We utilize this capability to develop a diagnostic test that probes for the presence of a pathogen’s DNA in body fluids.

We are developing an isothermal DNA-detection point-of-care diagnostic test that is equipment free and has a robust temperature range of operation. The DNA of a pathogen is detected with a CRISPR/Cas9 system using a guide RNA that is selected for repetitive sequences in the pathogen’s genome. The CRISPR/Cas9 system is coupled to an enzymatic DNA construct (DNAzyme), which produces a colorimetric readout when the pathogen’s DNA is detected. Rolling circle amplification is used to ensure an isothermal and sensitive reaction that does not require temperature control (in contrast to current DNA detection methods).

With this detection scheme, a simple color change, visible to the naked eye, will be produced when a pathogen’s DNA is present in a sample – making the test useful for resource-limited settings. The approach is broadly applicable and will enable self-diagnosis of a variety of pathogen-carried diseases including neglected tropical diseases such as leishmaniasis. Direct DNA detection is more reliable than immunological (antigen-antibody based) point-of-care tests as the effectiveness of the antibodies differ between populations, and antibodies persist after treatment, prevent the use of the current immunological tests for test-of-cure.

By adapting state-of-the-art technology into simple diagnostic tools, we are thus addressing one of the most pressing global health challenges that will benefit those who need it the most.