When a patient receives a kidney transplant, doctors carefully watch for signs of rejection in several ways, including biopsy. However, this procedure is invasive and can only detect problems at a late stage. Now researchers reporting in ACS ‘ Analytical Chemistry have developed a CRISPR-based test that can sensitive and non-invasively detect a biomarker of acute renal rejection in urine. It may someday help diagnose rejection earlier and without a biopsy.
Kidney transplant recipients must take immunosuppressive drugs for the rest of their lives to prevent their immune system from attacking the foreign organ. However, renal rejection can still occur, especially in the first few months after transplantation, which is known as acute rejection. Signs include increased serum creatinine levels and symptoms such as kidney pain and fever. Currently, the only way to definitively diagnose it is through a biopsy, but this procedure can only detect problems at a relatively late stage. Being able to diagnose renal rejection in a sensitive and non-invasive way at an early stage would allow doctors to start anti-rejection drugs sooner. Researchers previously found that high levels of a cytokine protein called CXCL9 in the urine of kidney transplant patients were a warning sign of rejection. But the current method of measuring CXCL9 (an enzyme immunoassay or ELISA) does not work very well in urine, which limits its sensitivity. Thus, Jonathan Dordick and his colleagues wanted to develop a more sensitive technique to non-invasively diagnose acute kidney rejection from urine.
The researchers based their detection method on CRISPR / Cas12a gene editing technology. In the presence of the CXCL9 protein, the CRISPR / Cas12a enzyme cuts a probe to produce a fluorescent signal. The researchers stimulated the fluorescent signal by attaching a DNA barcode that aggregates a large number of CRISPR / Cas12a molecules, and is then bound to an antibody that recognizes CXCL9. It is important to note that unlike other CRISPR-based detection methods, PCR amplification is not required, which makes it easy to adapt the method to a device that can be used in a doctor’s office or even at a patient’s home. When tested on urine samples from 11 kidney transplant patients, the new system accurately measured CXCL9 levels, with values ââvery similar to those of an ELISA. However, because the CRISPR immune system is about 7 times more sensitive than an ELISA, it may be able to detect kidney transplant rejection very early, the researchers say.
The authors acknowledge funding from the National Institutes of Health.
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Highly sensitive immuno-CRISPR assay for the detection of CXCL9
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