Abstract: SA-PO080

Development and Validation of 7-Plex Assay Panels to Measure Urine and Plasma Biomarkers Implicated in Prediction of CKD

Session Information

Category: Acute Kidney Injury

  • 003 AKI: Clinical and Translational

Authors

  • Sabbisetti, Venkata, Brigham and Women's Hospital, Boston, Massachusetts, United States
  • Christie, Emily, Brigham and Women's Hospital, Boston, Massachusetts, United States
  • Waikar, Sushrut S., Brigham and Women's Hospital, Boston, Massachusetts, United States
  • Bonventre, Joseph V., Brigham and Women's Hospital, Boston, Massachusetts, United States
Background

Biomarkers including TNFR1, TNFR2, suPAR, KIM-1, YKL 40, MCP-1 and BMP 7 have been shown to be associated with progression of kidney disease. We have developed and validated multiplex assays on the Meso Scale Discovery (MSD) platform to measure these 7 analytes simultaneously in both urine and plasma matrix.

Methods

We have developed a 7-Plex assay using MSD U-PLEX technology. Capture antibodies were biotinylated and conjugated to unique U-PLEX linkers and coupled to MSD 7-spot U-PLEX plates by incubating overnight at 4oC. Plates were coated with specific antibodies and then incubated with recombinant proteins, plasma or urine specimens from CKD patients for 1h followed by incubation with GOLD SULFO-TAGGED secondary antibodies for 1h. The amount of chemiluminescence in each well was measured using MSD SQ120 instrument.

Results

The MSD 7-plex assay demonstrated excellent linearity of dilution and spike recovery with no interference of various substances including glucose, bilirubin and albumin in both urine and plasma matrix. The intra and inter-assay coefficient of variation was below 15%.

Conclusion

We have developed a robust and reliable 7-plex assay on MSD platform to measure biomarkers implicated as potentially predictive of progression of kidney disease in plasma and urine specimens.

Funding

  • NIDDK Support