Table 3.

Evaluation of antibody molecular properties that are most strongly correlated with non-specific interactions in physiological conditions (PBS, pH 7.4) and self-association in formulation conditions (10 mM histidine, pH 6). The sequence-based charge features were calculated as described in Table 1. The ELISA/PSP results are for the average measurements from the four assays. The structure-based features were evaluated from Fv homology models using Molecular Operating Environment software.

Molecular Property		Region	Descriptors	ELISA #1 (pH 7.4)		ELISA #2 (pH 7.4)		PSP #1 SMP (pH 7.4)		PSP #2 OVA (pH 7.4)		ELISA/PSP (pH 7.4)		kD, mL/g (pH 6)
Molecular Property		Region	Spearman’s correlation	ρ	p-value	ρ	p-value	ρ	p-value	ρ	p-value	ρ	p-value	ρ	p-value
Sequence Based	Electrostatic	HCDR3	Net charge	0.34	^*	0.25	ns	0.09	ns	0.34	^*	0.26	ns	0.12	ns
		CDR	Net charge	0.80	^****	0.76	^****	0.29	ns	0.55	^***	0.69	^****	0.49	^*
		V_L	Net charge	0.78	^****	0.79	^****	0.50	^**	0.46	^**	0.87	^****	0.76	^***
		V_H	Net charge	0.77	^****	0.81	^****	0.32	^*	0.59	^****	0.68	^****	0.62	^*
		Fv	Net charge	0.88	^****	0.89	^****	0.42	^**	0.55	^***	0.88	^****	0.86	^****
		Fv	Cross charge (V_H xV_L)	0.82	^****	0.75	^****	0.53	^**	0.53	^***	0.86	^****	0.63	^*
		Fv	pI	0.86	^****	0.83	^****	0.48	^*	0.56	^***	0.88	^****	0.84	^****
		IgG	pI	0.85	^****	0.84	^****	0.48	^*	0.56	^***	0.89	^****	0.84	^****
Structure Based (Fv)	Stability	Fv	Buried SA (V_H/V_L interface); Å²	−0.53	^**	−0.50	^**	0.26	ns	0.14	ns	−0.32	^*	−0.18	ns
	Stability	Fv	Interaction energy (V_H/V_L interface); Å²	0.59	^***	0.56	^**	−0.15	ns	−0.07	ns	0.40	^**	0.22	ns
	Electrostatic	Fv	Hydrophilic SA; Å²	0.76	^****	0.73	^****	0.04	ns	0.16	ns	0.60	^****	0.52	^*
		Fv	Positive patch SA; Å²	0.82	^****	0.77	^****	0.37	^*	0.38	^*	0.82	^****	0.71	^**
		Fv	Negative patch SA; Å²	−0.09	ns	0.01	ns	0.02	ns	0.11	ns	−0.16	ns	−0.20	ns
		CDR	Positive patch SA near CDR; Å²	0.84	^****	0.82	^****	0.28	ns	0.40	^**	0.75	^****	0.60	^*
		CDR	Negative patch SA near CDR; Å2	−0.08	ns	0.01	ns	0.24	ns	0.23	ns	−0.01	ns	−0.02	ns
		Fv	Dipole moment (Debye)	0.26	ns	0.30	Ns	0.27	ns	0.48	^**	0.22	ns	−0.06	ns
		CDR	Net charge	0.72	^****	0.67	^****	−0.12	ns	0.15	ns	0.48	^**	0.48	^*
		Fv	Net charge	0.81	^****	0.79	^****	0.57	^****	0.63	^****	0.88	^****	0.75	^***
		Fv	pI	0.84	^****	0.81	^****	0.53	^***	0.62	^****	0.89	^****	0.82	^****
	Hydrophobic	Fv	Zeta potential (mV)	0.81	^****	0.79	^****	0.57	^****	0.63	^****	0.88	^****	0.75	^***
		Fv	Hydrophobic SA; Å²	−0.49	^**	−0.48	^*	−0.11	ns	−0.05	ns	−0.48	^**	−0.65	^*
		Fv	Hydrophobic SA; Å²	−0.35	^*	−0.37	^*	−0.20	ns	−0.36	^*	−0.32	^*	−0.20	ns
		CDR	Hydrophobic patch SA near CDR; Å²	−0.67	^****	−0.65	^****	−0.47	^**	−0.42	^**	−0.77	^****	−0.69	^**
		Fv	Hydrophobic moment	0.56	^**	0.51	^*	−0.27	ns	−0.14	ns	0.31	^*	0.31	ns
		Fv	Hydrophobic imbalance	0.51	^**	0.48	^*	0.15	ns	0.00	ns	0.57	^****	0.42	^*

Spearman’s ρ and p-values are reported, the latter which are indicated as values <0.0001 (****), <0.001 (***), <0.01 (**), 0.01 to 0.05 (*), and >0.05 (ns). Surface area is referred to as SA.