Label-free live-cell imaging and endpoint cell viability data obtained from exposing the patient derived glioblastoma cell line U3118 (see related article below), on all combinations of the CUSP9v3 protocol from order 5 to 9. The standard-of-care drug, temozolomide, was used as add-on to all combinations. There are 4 .zip files containing all the imaging data, one for each 384-well plate used. Every .zip file contains 5544 grayscale images of size 800x992 pixels and every experimental well consists of a temporal sequence of 18 such images. The experiment was performed in quadruplicate and the associated plate layouts used for acoustic liquid dispension are also included as .xlsx files. The .log file contains the cell viability data for all plates.