Protocol for the generation and automated confocal imaging of whole multi-cellular tumor spheroids
Publication Name
STAR Protocols
Abstract
Multi-cellular tumor spheroids (MCTS) have found widespread use in pre-clinical research. However, their complex three-dimensional structure makes immunofluorescent staining and imaging challenging. Here, we present a protocol for whole spheroid staining and automated imaging using laser-scanning confocal microscopy. We describe steps for cell culture, seeding of spheroids and transfer of MCTS, and adhesion to Ibidi chamber slides. We then detail fixation, immunofluorescent staining based on optimized reagent concentrations and incubation times, and confocal imaging facilitated by glycerol-based optical clearing.
Open Access Status
This publication may be available as open access
Volume
4
Issue
2
Article Number
102331
Funding Number
APP1181179
Funding Sponsor
FightMND