Journal: Nature Methods

Article Title: MISpheroID: a knowledgebase and transparency tool for minimum information in spheroid identity

doi: 10.1038/s41592-021-01291-4

Figure Lengend Snippet: Representative microscopy images show ethidium homodimer I stained (red if cell is dead) spheroids of all evaluated cell lines and patient-derived sarcoma cells (cell types indicated on the left) cultured in different media types (indicated on the top) (scale bars 200 µm). Each experiment was repeated independently at least 3 times with 8 technical replicates per experiment, with similar results. Media types are ranked from higher nutrient (left) to lower nutrient (right) richness. Intense staining in the spheroid center is indicative of necrotic core. Next to cell death, images give an indication of circularity and size.

Article Snippet: For the comparison of the liquid overlay method and the hanging drop method, spheroids were cultured for 3 d. A total of 60 μl medium per well was replaced with 10 μM Ethidium homodimer I solution (cat. no. 300519, Santa Cruz) in PBS supplemented with Ca 2+ and Mg 2+ .

Techniques: Microscopy, Staining, Derivative Assay, Cell Culture

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