We have used a quick and dirty protocol as below (based on the protocol of the DECIPHER shRNA library from Cellecta) for endothelial cells transduced with lentiviral vectors and puromycin marker, which has worked fine for us:
Day
1 Seed 2000-5000 cells per 96 well in medium
2 Infect cells with indicated viral dilutions in medium with 5 μg/ml Polybrene
3 Replace medium with fresh medium supplemented with 2 μg/ml (or equivalent 99.9% killing concentration of) Puromycin
4
5 Viability assay (resazurin)
Your readout should look something like this (see attachment)