DirectBio is pioneering a new approach to molecular diagnostics based on CRISPR enzymes.

We harness the speed, specificity, and sensitivity of natural and AI-engineered CRISPR enzymes to create diagnostic assays that are simpler and more scalable than conventional PCR due to the need for fewer reagents, no RNA or DNA extraction, and no amplification or reverse transcription.

Direct Detection RNA Panels

Beginning with Cas13a, which uses a guide RNA to recognize a sequence of single-stranded target RNA and turn on indiscriminate RNase activity, we are developing direct detection RNA panels for both laboratory medicine and point-of-care applications. Cas13a generates fluorescence upon target RNA recognition through cleavage of fluorophore-quencher pairs tethered by single-stranded RNA that are added along with the enzyme and guide RNAs to a sample such as saliva, blood, or a nasal swab that has been minimally processed by heat and RNase inhibitors. The fluorescent signal generated by Cas13a is in direct proportion to the number of copies of target in the reaction, resulting in quantitative assays that can be read on existing equipment from multiple diagnostic companies, much like PCR reagents. Confining these CRISPR reactions to droplets or microwells enables sensitivity at or below 1 copy of target per microliter of reaction, with further improvements likely from optimized enzymes and reporter molecules.

Kinetic Barcoding

In addition to easily changing the guide RNA sequence to recognize different RNA targets, we have developed modifications to the guides that allow us to control enzymatic rates for undiluted multiplexing through kinetic barcoding and to detect the presence of proteins and metabolites from the same samples using Cas enzymes. We envision a CRISPR-based future for diagnostics with point-of-care multi-analyte assays that combine the diagnostic power of multiple biomarkers from individual samples to efficiently detect and monitor patient health.

Altering the Enzyme

In addition to the above, DirectBio is altering the enzyme and guides to target non-nucleotide biomarkers and to provide multiplexed, multi-biomarker type, syndromic panels. This ability to detect molecules other than RNA and DNA builds on the guide modifications that enable kinetic barcoding.

The innovation of DirectBio has the following advantages:

  • Multiplexed, multi-modality (multi-biomarker type) syndromic or disease specific panels
  • Simplicity to handle at patient site device for Point of Care (POC) applications, as the over 20-year-old existing PCR is difficult and complicated
  • The DirectBio technology works on some of the existing PCR platforms, POC and central labe clinical analyzers.
  • True Multiplexing. Multiplexing not by dividing the sample but by multiplexing in one solution; One-Sample Multiplexing
  • In contrast to other methods, the DirectBio technology allows for quantifiable results
  • The DirectBio technology is less error prone as it requires no amplification steps compared to conventional PCR technologies
  • The DirectBio technology has similar and better sensitivity compared to conventional PCR technologies.
  • The DirectBio technology is faster as it requires no amplification steps compared to conventional PCR technologies
  • Due to all reasons above, the diagnostics becomes cheaper compared to conventional PCR technologies
  • The DirectBio technology allows physicians to do all diagnostic testing on site in their own practice and receive additional income from paying insurances, which would otherwise have gone to central labs.