Journal: Oncotarget

Article Title: Molecular imaging reveals biodistribution of P-cadherin LP-DART bispecific and trafficking of adoptively transferred T cells in mouse xenograft model

doi: 10.18632/oncotarget.27544

Figure Lengend Snippet: ELISA based assay was used to evaluate the effect of VT680 labeling on binding of P-cadherin LP-DART to human P-cadherin and human CD3. The proteins were coated on to the plates and incubated with serial dilutions of P-cadherin LP-DART-VT680 or Control LP-DART-VT680 for 1 hr at 37°C. The bound P-cadherin LP-DART was quantified using IgG-HRP conjugate followed by calorimetric quantitation. ( A ) The labeling of VT680 to P-cadherin LP-DART had a DOL dependent effect on binding to human P-cadherin and Control LP-DART had no binding to human P-cadherin. ( B ) The labeling of VT680 to P-cadherin LP-DART and Control LP-DART affected binding to human CD3. P-cadherin LP-DART retained moderate binding even at a DOL of 2.0, whereas Control LP-DART lost its binding to human CD3 protein. ( C ) CTL Assay: Firefly luciferase expressing HCT116 cells and expanded human CD3+ T lymphocytes were co-incubated with increasing concentrations of P-cadherin LP-DART with different DOL of VT680. After 24 hr the remaining viable cells were quantified by measuring the luciferase activity. The relative cytotoxicity observed at various concentrations of P-cadherin LP-DART-VT680 was plotted against the PBS treated samples. VT680 conjugation decreased the cytotoxic ability in a DOL dependent manner.

Article Snippet: EC50 was determined by non-linear curve fit of percent cytotoxicity using GraphPad Prism 7.04 software (GraphPad Software).

Techniques: Enzyme-linked Immunosorbent Assay, Labeling, Binding Assay, Incubation, Control, Quantitation Assay, CTL Assay, Luciferase, Expressing, Activity Assay, Conjugation Assay