Isolation and light chain shuffling of a Plasmodium falciparum AMA1-specific human monoclonal antibody with growth inhibitory activity

Table 4 Expression levels and affinity constants assessed by SPR spectroscopy

	OcheA-9	OcheA-16	OcheA-17	OcheA-17B
Expression level (mg/kg FLM)	150	140	440	110
AMA1 DiCo1 K_D (nM)	–	–	213	43
AMA1 DiCo2 K_D (nM)	110	–	160	1750
AMA1 DiCo3 K_D (nM)	68	55	61	1.7
Pf3D7 AMA1 K_D (nM)	–	–	–	176

To calculate expression levels the concentrations of recombinant antibodies OcheA-9, OcheA-16, OcheA-17 and OcheA-17B in N. benthamiana plant extracts were assessed measuring the capture level on a protein A surface in relation to an internal antibody standard using BiacoreT200. For affinity kinetic measurements purified recombinant antibodies OcheA, OcheA-9, OcheA-16, OcheA-17 and OcheA-17B were captured on a protein A surface (110, 110, 130, 140, 120 RU respectively) and the antigens were injected successively in concentrations of 360 nM, 180 nM and 90 nM for 180 s (30 µL/min). The surface was washed with HBS-EP buffer and regenerated with 30 mM HCl between measurements. Data were evaluated using T200EvaluationSoftware3.0 using the two state reaction binding model. “–”: not determinable

ISSN: 1475-2875