Table 1. 2D screening platforms for cartilage.

Authors	Cell type(s)	Model	Chondrogenic medium?	Molecules/parameters tested	Culture period	Outcomes
Johnson et al.(2012) 29	Human BMSC	384-well format. Cells seeded in monolayer and molecules added to medium.	No	22,000 structurally diverse, heterocyclic, drug-like molecules (5 μM). KGN hit.	4 days	Presence of chondrogenic nodules with rhodamine B staining and light microscopy
Rape et al. (2015)44	Human glioblastoma cells and ASC	Combinatorial hydrogels. HA gels with stiffness and fibronectin density gradients cured onto glass slides. Cells seeded on top.	No	Different stiffnesses and fibronectin concentrations	2 days (glioblastoma) and 7 days (hASC)	Cancer model: miR18a expression via fluorescence assay. Adipogenic/osteogenic model: Oil Red O and NBT/BCIP staining.
Floren and Tan (2015) 42	Rodent BMSC/PASMC	ECM microarray. Electrospun PEGDM and PEO deposited onto glass slides and photopolymerised. Spotted with different ECM proteins. Cells seeded on top	No	Collagen I, collagen III, collagen IV, laminin, fibronectin, elastin	24 hours	Cell adhesion and spreading with DAPI and phalloidin. ICC (PECAM-1 vascular marker). Imaged with automated confocal microscopy
Le et al. (2015) 12	ATDC5 chondrogenic cell line	96-well format. Cells seeded in monolayer and molecules added to medium.	Yes	LOPAC library of 1280 pharmaceutically active compounds	9 days	Total collagen via fluorescent collagen probe assay.
Choi et al. (2016)33	Human ASC	60 mm plates. Cells seeded in monolayer and molecules added to medium.	No	In-house protein kinase inhibitor library of sulphonamides (1 μM and 10 μM). Compound 6 hit.	11 days	Aggrecan expression via ELISA
Tong et al. (2016)45	Human fibroblasts	Combinatorial PEG hydrogels with mechanical and ligand density gradients formed on glass slides and cells seeded on top.	No	Different stiffnesses and RGD binding densities	24 hours	Cell adhesion and morphology, cytoskeleton structure and spreading/elongation all via staining and microscopy
Sharma et al. (2017) 7	Various, including bovine chondrocytes	PEG hydrogels electrospun onto glass slide and UV cured, then peptide microarray deposited on top via array spotting system. Cells (mono- and co-cultures) seeded on top.	Yes (chondrocytes only)	Multiple peptide motifs and concentrations	24 hours	Cell adhesion and morphology and cytoskeleton structure via staining and confocal microscopy
Ming et al. (2018) 8	Rodent chondrocytes	Microfluidic device used to create concentration gradient of molecule. 8 concentrations of medium/drug directed to cells cultured in monolayer in 8 downstream chambers.	No	Resveratrol (0-200 μM)	3 days	Proliferation via cell counting via light microscopy
Gobaa et al. (2018) 43	Human BMSC	2016-well format, combinatorial protein array. Proteins deposited onto thin layer of PEG hydrogel via robotic liquid handling system and cells seeded on top.	No	Wnt3a, Wnt5a, DKK1, BMP2, DLL4, Jag, DLK1, NCAM, GDF8, CCL2, laminin Different PEG stiffnesses	11 days	Proliferation with DAPI and phalloidin and adipogenic differentiation with Nile Red staining. Imaged with automated microscopy
Deshmukh et al. (2018) 10	TCF/LEF reporter cell line and human BMSC	Molecules adhered to multi-well screening plates via robotic liquid handling system and cell seeded on top.	Yes (BMSC only)	Wnt pathway inhibitors: SM04690, FH535, IWR-1, ICG001, iCRT14, KY02111, CX.4945	2 days (TCF/LEF reporters) 5 days (hBMSC)	Luciferase activity (TCF/LEF reporters) Presence of chondrogenic nodules with rhodamine B staining and automated imaging system (hBMSC)
Nogueira-Recalde (2019) 9	T/C28a2 chondrocyte cell line	384-well format. Cells seeded in monolayer. Aged/senescent model induced with IL-6. Automated cell dispensing and liquid handling.	No	Prestwick chemical library of 1120 approved drugs	Not specified	Senescence-associated-β-galactosidase activity via ImaGene Green™ C12FDG lacZ Gene Expression Kit. Autophagy levels via LC3 reporter. Imaged with Operetta® High Content Screening system.
Shi et al. (2019) 11	Murine chondrogenic cell line	96-well format. Cells seeded in monolayer and molecules added to medium.	No	Library of 2320 natural and synthetic small compounds (10 μM). BNTA hit.	5 days	Alcian Blue stain for proteoglycans assessed via light microscopy

ASC = adipose-derived stem cell. BMSC = bone marrow stromal cell. ECM = extracellular matrix. ELISA = enzyme-linked immunosorbent assay. HA = hyaluronic acid. ICC = immunocytochemistry. PEG = poly(ethytlene glycol). PEO = poly(ethylene oxide). PASMC = pulmonary arterial smooth muscle cells. RGD = Arg-Gly-Asp tri-peptide