Fig. 4: BsAb formation can be detected in raw reaction samples via bridging-ELISA and be quantified by SEC.

a Bridging-ELISA and detection of functional bsAbs in raw reaction mixtures without prior purification. Red lines represent hole–knob combinations, blue lines the knob–hole combinations of the respective target antibodies.The green line in the upper right panel is a conventionally produced bsAb control. Additional traces are negative controls (raw reaction mixtures of control antibodies that do not match the antigens used in the respective bridging ELISA). b Bridging ELISA indicates that productive exchange requires reduction for initiation. SEC (see also left panel of Fig. 3b) quantification determines TCEP concentrations that enable effective initiation of exchange (TCEP dependent formation of exchange products after 1 h 37 °C monitored by analytical SEC).