Journal: ACS Applied Materials & Interfaces

Article Title: Cryopreservation of Liver-Cell Spheroids with Macromolecular Cryoprotectants

doi: 10.1021/acsami.2c18288

Figure Lengend Snippet: Post-thaw (24 h) recovery of cryopreserved HepG2 spheroids. Percentage viability of spheroids (A) frozen directly in agar molds and stored at −80 °C for 24 h; (B) frozen suspended in cryovials and stored at −80 °C for 24 h; and (C) frozen in cryovials and stored above liquid nitrogen for 3 days. Data are presented as mean % viability relative to pre-frozen spheroids ± SEM from five independent repeats, determined by the WST-1 assay. (D) Confocal microscopy of spheroids before and after thawing in DMSO (10%) with and without supplementation with 20 mg mL –1 polyampholyte. Living cells are labeled green (calcein-AM, intact membrane), dead cells are labeled red (EthD-III, damaged membrane), and the nuclei of cells with intact membranes were stained with Hoechst 33342 solution (20 mM). Scale bar = 100 μm.

Article Snippet: Live/Dead viability assay kit Ethidium homodimer-1 (EI) and calcein-AM (L3224), Corning 96-well white polystyrene microplates (10022561), trypsin (0.25%) and EDTA phenol red (500 mL) (25200072), Invitrogen ActinGreen 488 ReadyProbe reagent (R37110), antibiotic–antimycotic solution 100× (15240062), and cryovials were purchased from Thermo Fisher (Loughborough, UK).

Techniques: WST-1 Assay, Confocal Microscopy, Labeling, Staining