PROJECT SUMMARY For the 786,000 people in the United States living with end stage renal disease (ESRD), the treatment of choice is kidney transplantation. While transplant life has subtly increased over the last 15 years, ten-year graft survival rates are only 28% for deceased-donor and 46% for living-donor grafts. The primary cause of graft failure after the first year is antibody-mediated rejection (AMR), the detrimental impacts of which are compounded by concurrent fibrosis. Thus, there is an urgent yet unmet need to prolong graft life by remediating AMR prior to extensive fibrotic changes and other severe, irreversible graft damage. Therapeutic interventions to remediate AMR are not FDA approved or clinically standardized due to their inconsistent and often suboptimal performances. Promising treatment protocols are in development, but their translation to clinical trials will require frequent monitoring of AMR response that cannot be supported by the current evaluation standard, invasive biopsy, due to its associated safety risks. A safer alternative involves tracking surrogate serum and urine biomarkers, but these markers are nonspecific with unclear diagnostic thresholds, and measurable fluctuations in their values may occur well after underlying pathological changes to the allograft. Another safe alternative to biopsy is imaging, most often ultrasound, which also indicates AMR nonspecifically. The lack of a noninvasive and specific method for detecting AMR and routinely monitoring its response to treatment in renal transplant patients represents a major gap in prolonging graft life. To fill this gap, our group has spent the first five-year cycle of this grant award developing noninvasive Viscoelastic Response (VisR) ultrasound for monitoring renal transplant status. Our research has demonstrated that in vivo VisR assessments of elastic and viscous differences across the medulla and cortex, as well as medullary and cortical elastic and viscous anisotropy, delineate renal inflammation and fibrosis in a pig model. Further, in a pilot clinical feasibility study, our research has shown that in vivo VisR differentiates biopsy-confirmed tubulointerstitial fibrosis in the allografts of human patients imaged serially from time of implantation. The success of our investigations thus far motivates further advancement of VisR technology and its extension to noninvasively detecting and monitoring AMR. Specifically, we will augment the VisR methods developed in the first funding cycle with a new approach to anisotropy measurement (angle interpolation) and a new imaging method (Double Profile Intersection (DoPIo) ultrasound) to improve the sensitivity and specificity of detecting inflammation, a hallmark of AMR. We will apply the advanced VisR and DoPIo methods to identifying AMR and monitoring its response to treatment in human renal transplant patients. We hypothesize that advanced VisR and DoPIo ultrasound delineate biopsy-confirmed inflammation assoc...