Fig 1: Cells were isolated from dog NP tissue using different methods. Tissues digested with collagenase alone (C) compared with collagenase with added pronase (C + P) after a 4 or 16 h digestion period. Viability (A) and live number of NP cells/g tissue (B) isolated from CD dog NP tissue following different collagenase type II digestion times (4 or 16 h) with/without 30 min of pronase predigestion. Gene expression for ACAN (C), COL2A1 (D), BAX (E), BCL2 (F), COL1A1 (G), KRT8 (H), and KRT19 (I) normalized for the mean expression of reference genes GAPDH, HPRT, and RPL19 immediately following extraction of NP cells from dog tissue. Digestion methods: T4c = 62.5 U/ml collagenase type II for 4 h; T4c + p = 5.25 U/ml pronase for 30 min followed by 62.5 U/ml collagenase type II for 4 h; T16c = 62.5 U/ml collagenase type II for 16 h; T16c + p = 5.25 U/ml pronase (30 min) followed by 62.5 U/ml collagenase type II for 16 h. Viability analysis was tested using Friedman test for paired data with Dunns multi-comparison post hoc testing. Live cell number and gene expression analysis was analyzed with ANOVA with Tukey's post hoc test *P < 0.05. n = 3 dogs (A and B), in triplicates (D–I).