The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.

The cytotoxicity of hIMB1636-MMAE and free MMAE was evaluated by the Cell Counting Kit-8 (CCK-8) assay (Dojindo, Kyushu, Japan). Cells were plated at 2 × 103 to 8 × 103 cells/well in 100 uL culture medium into 96-well plates. Following an overnight incubation at 37 °C, the cells were cultured continuously for 48 h in 100 uL of medium containing various concentrations of drugs (100, 10, 1, 0.1, 0.01, and 0.001 nM). Then, the medium was replaced with 100 uL of fresh medium containing 10% CCK-8 reagent and incubated for 1 h. The absorbance was measured at 450 nm by a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, LLC, USA). The half-maximal inhibitory concentration (IC50) of the samples was calculated by Prism GraphPad 6.0 (GraphPad Software, San Diego, CA). The data are presented as the mean ± SD for three independent experiments.