Fig. 4

Competition ELISAs to test for overlapping epitopes recognized by the scFvs binding ALS selective TDP-43 variants. Competition capture ELISAs were utilized to determine if the scFvs bound similar epitopes. Samples of homogenized ALS, FTD and healthy brain tissue (motor cortex) were pre-incubated with the ALS-TDP6, ALS-TDP13, ALS-TDP15, ALS-TDP18 and AD-TDP3 scFvs individually for ~1 h. These samples were then utilized in capture ELISAs with the same five scFvs immobilized to the wells: a ALS-TDP6, b ALS-TDP13, c ALS-TDP15, d ALS-TDP18 and e AD-TDP3. Bound phages were detected with an anti-M13-HRP antibody and signal intensities calculated based on the difference between the ALS and FTD samples. All the scFvs produced similar binding signal strengths with or without the presence of the competitive scFvs. One-way ANOVA with LSD post hoc analysis supported the competitor scFvs’ inability to block the recognized antigenic site of our capture scFvs since no significant differences were detected between its presence and the no competition sample. Overall, these results demonstrated recognition of multiple TDP-43 variants by our scFvs