Table 1.

Summary of single-cell isolation protocols for nucleus pulposus in reported studies using scRNA-seq technique

Reference	Classification of IVD	Species	Treatment	Dead cell removal	Profiling cellular types
Gan Y, et al. [10]	Pfirrmann grade I	Human	TrypLE Express for 30 min, 0.2% pronase (Sigma-Aldrich, USA) for 60 min, and 0.012% collagenase II (Sigma-Aldrich, USA) for 2 ~ 4 h, sequentially	Yes	Chondrocyte, Notochord, Nucleus pulposus progenitor cells, Stromal cell, Pericyte, Endothelial cell and blood cell
Fernandes LM, et al. [11]	Thompson grade 1 or 2	Human	Pronase for 1 h and then collagenase P overnight	No Culture cells for scRNA-seq	Cultured nucleus pulposus cells
Calió M, et al. [12]		Bovine	0.19% Pronase for 1 h and then 260 IU/mg Collagenase II overnight	No	Nucleus pulposus cells, uncharacterized cell subset, monocyte derived dendritic cells
Zhang Y, et al. [13]	Pfirrmann grade I-IV	Human	GEXSCOPETM Tissue Dissociation Solution for 15 min. (Included in the GEXSCOPE® Single Cell RNA library Kit)	No	Chondrocytes, macrophages and endothelial cells
Ji Tu, et al. [14]	Pfirrmann grade II-V	Human	500 U/mL collagenase I, 150 U/mL collagenase II, 50 U/mL collagenase IV, 0.1 mg/ml hyaluronidase and 30 U/mL DNaseI for 95 min	Yes	Neucleus pulposus cells, natural killer cells, monocytes, and T cells