A Bioinspired Immunostimulatory System for Inducing Powerful Antitumor Immune Function by Directly Causing Plasma Membrane Rupture

Preparation of AMDs. 125 μM Ac4ManNAz were incubated with Raw 264.7 (~ 107 cells/dish) for 24, 48, and 72 h, followed by gentle washing with PBS or complete medium to produce Raw 264.7-Ac4ManNAz. Subsequently, Raw 264.7-Ac4ManNAz (~ 107 cells) were resuspended in PBS containing Alkynyl-Ce6(0.1 μM), ascorbic acid (0.2 μM) and CuSO4 (0.2 μM) and incubated for 1h at 37 °C. Then centrifuged (1000 rpm, 3 min) and washed 3 times with PBS to obtain precursor of AMDs. precursor of AMDs (~ 107 cells/tube, 1.0 ml) was immersed in liquid nitrogen, 24 h later, the frozen tubes were thawed at 37°C, centrifuged at 1500 rpm for 3 min, washed and resuspended by PBS to generate the AMDs (5 x 106 units/tube, 1.0 ml).