Doctors monitor kidney transplant recipients for signs of rejection in several ways, including biopsy. These tests are invasive; therefore, they can only detect problems at a late stage.
A CRISPR-based method could help diagnose kidney transplant rejection early and non-invasively. Image Credit: phugunfire/Shutterstock.com.
Researchers have now created an assay based on CRISPR that can detect a biomarker of acute kidney rejection in urine using a highly sensitive and non-invasive method. In the future, this may make rejections easier to diagnose without an invasive biopsy. The study was published in ACS’ Analytical Chemistry.
Immunosuppressive drugs must be taken for the rest of the recipient’s life to prevent their immune system from attacking the transplanted organ. It is still possible for kidney rejection to occur, especially during the first few months following transplantation, which is called acute rejection. Symptoms include kidney pain and fever as well as abnormal serum creatinine levels.
A biopsy is the only way to diagnose it definitively, but this procedure can detect the problem at relatively late stages. It would be beneficial if kidney rejection could be detected non-invasively at an early stage, allowing doctors to begin anti-rejection medication sooner.
In previous studies, researchers found that the presence of high levels of a cytokine called CXCL9 in the urine of kidney transplant patients was an early indication of rejection.
However, the present method for measuring CXCL9 (an enzyme-linked immunosorbent assay, or ELISA) does not work well in urine, thus limiting its sensitivity. To better diagnose acute kidney rejection non-invasively, Jonathan Dordick and colleagues wanted to develop a more sensitive method.
The detection method developed was based on CRISPR/Cas12a gene-editing technology. In the presence of the CXCL9 protein, the CRISPR/Cas12a enzyme incise a probe to produce a fluorescent signal. To boost the fluorescent signal, researchers attached a DNA barcode that aggregates CRISPR/Cas12a molecules and is bound to an antibody specific for CXCL9.
As a result, unlike other CRISPR-based detection methods, this one does not require PCR amplification, which makes it easier to adapt to a device that can be used in a doctor’s office or even at home. Testing on urine samples from 11 kidney transplant patients, the new system accurately determined CXCL9 levels, resulting in values similar to an ELISA test.
According to the researchers, however, by being seven times more sensitive than ELISA, immuno-CRISPR could be used to identify kidney transplant rejection at a very early stage.
Lee, I., et al. (2021) Highly Sensitive Immuno-CRISPR Assay for CXCL9 Detection. Analytical Chemistry. doi.org/10.1021/acs.analchem.1c03705.