Uman Type NF-L Antibodies Are Effective Reagents for the Imaging of Neurodegeneration

Abstract

AbstractRecent work shows that certain immunological assays for the neurofilament light chain NF-L detect informative signals in the CSF and blood of human and animals affected by a variety of CNS injury and disease states. Much of this work has been performed using two mouse monoclonal antibodies to NF-L, UD1 and UD2, also known as 2.1 and 47.3 respectively. These are the essential components of the Uman Diagnostics NF-Light™ ELISA kit, the Quanterix Simoa™ bead based NF-L assay and others. We show here that the antibodies bind to neighboring epitopes in a short, conserved and unusual peptide in the NF-L “rod” Coil 2 region. We also describe a surprising and useful feature of Uman and similar reagents. While other well characterized NF-L antibodies show robust staining of countless cells and processes in CNS sections from healthy rats, both Uman antibodies reveal only a minor subset of presumably spontaneously degenerating or degenerated neurons and their processes. However following experimental mid-cervical injuries to rat spinal cord both Uman antibodies recognize numerous profiles in tissue sections. The Uman positive material was associated with fiber tracts expected to be damaged by the injury administered and the profiles had the swollen, beaded, discontinuous and sinusoidal morphology expected for degenerating and degenerated processes. We also found that several antibodies to the C terminal “tail” region of NF-L stain undamaged axonal profiles but fail to recognize the Uman positive material. The unmasking of the Uman epitopes and the loss of the NF-L tail epitopes can be mimicked by treating sections from healthy animals with proteases suggesting that the immunological changes we have discovered are due to neurodegeneration induced proteolysis. We have also generated a novel panel of monoclonal and polyclonal antibody reagents directed against the region of NF-L including the Uman epitopes which have staining properties identical to the Uman reagents. Using these we show that the NF-L region to which the Uman reagents bind contains further hidden epitopes distinct from those recognized by the two Uman reagents. We speculate that the Uman type epitopes are part of a binding region important for higher order neurofilament assembly. The work provides important insights into the properties of the NF-L biomarker, describes novel and useful properties of Uman type and NF-L tail binding antibodies and provides a hypothesis relevant to further understanding of neurofilament assembly.