Abstract: SA-PO544
Developing a Highly Sensitive Meso Scale Discovery (MSD) SLIT Immunoassay for Chronic Kidney and Eye Disease
Session Information
- Bioengineering
October 26, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Bioengineering
- 400 Bioengineering
Authors
- Liaw, Easton J., Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Kumar, Sudhir, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Fan, Xueping, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Huynh, Courtney, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Schmidt, Insa Marie, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Shoushtari, Sara Itzel, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Sheikh, Zahra, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Subramanian, Manju L., Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Waikar, Sushrut S., Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
- Lu, Weining, Boston University Chobanian & Avedisian School of Medicine, Boston, Massachusetts, United States
Background
Both kidney failure and blindness are common complications of poorly controlled diabetes mellitus, which afflicts over 150 million people around the world. The kidney and eye share several structural similarities and developmental pathways that may account for their shared susceptibility as end-organ complications of diabetes. SLIT is a ligand for ROBO receptors. SLIT/ROBO pathway plays essential roles in kidney and eye development. Published ELISA results showed upregulation of SLIT protein expression in both chronic kidney and eye diseases. Recently, a novel SLIT inhibitor has advanced to a phase II clinical trial for proteinuric kidney disease. However, the low sensitivity and dynamic range of the ELISA assay have made the SLIT immunoassay results unreproducible.
Methods
We developed a highly sensitive Meso Scale Discovery (MSD) immunoassay using an electrochemiluminescence detection technique with a greater dynamic range that may replace previously developed ELISAs for SLIT detection. In a process similar to a sandwich ELISA assay, we detected and quantified standard SLIT recombinant protein and unknown SLIT concentration in human and mouse samples using the MESO QuickPlex SQ120 instrument.
Results
Our data showed a greater dynamic range and more precise and accurate readings on a lower concentration of SLIT using the MSD assay (LLoD=5 pg/ml, ULoD=20,000 pg/ml) compared to regular ELISA (LLoD=1,000 pg/ml, ULoD=50,000 pg/ml). We also found the optimal concentrations of the capture and detection antibodies that best suit the SLIT measurements. Besides the newly developed SLIT MSD assay able to detect low protein levels, we also discovered 7-fold higher SLIT levels in vitreous humor compared to blood samples in humans and an 80% serum SLIT level reduction in mice treated with a SLIT inhibitor.
Conclusion
We have developed a highly sensitive MSD assay across 4 orders of magnitude for SLIT in humans and mice. This assay may be used in clinical trials as a target engagement biomarker for SLIT inhibitors and in preclinical animal models of diabetic kidney and eye diseases.
Funding
- NIDDK Support – Pfizer