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. 2024 Jan 5;13(1):5. doi: 10.3390/antib13010005

Figure 3.

Figure 3

ELISA and FACS assays to characterize hybridoma supernatant properties. (A) Coomassie staining of purified hACE2 protein and WT RBD-6xHis in reducing and non-reducing conditions. Both produced proteins have been used in subsequent ELISAs. MW RBD-6xHis 26 KDa, MW hACE2 145 KDa. (B) SEC analysis of the hACE2 (upper) and WT RBD-6xHis (lower) to evaluate purity of the produced proteins. (C) Schematic representation of the competition ELISA assay performed on selected hybridoma supernatants. (D) Competitive ELISA, results are shown as A.U. (405 nm). High competing mAbs display low IC50 values. (E) FACS assay on hACE2-expressing Vero cells, using hybridomas culture media in three serial dilutions (1:2, 1:20, 1:200). Antibodies neutralizing the binding display 0% binding percentage.