Comparison of expression and activity of TGM2 in SB623 and MSC; its functional analysis in ECM using siRNA. (A) Expression levels of TGM2 normalized to GAP were determined by using qRT-PCR in SB623/MSC pairs from several donors. Levels in SB623 cells were expressed relative to levels in parental MSCs, which were set on 1. (B) TGM2-crosslinking activity was measured by amounts of biotinylated cadaverine incorporated into PLL in the presence of SB623 or MSC cell lysates. The activity was then normalized to the total protein, and expressed compared with the parental MSCs, where the values were set on 1. (C) TGM2 was detected in ECM of MSCs and SB623 by using immunoblotting. The TGM2 signal was quantified densitometrically and normalized to the total ECM protein per lane. The total ECM protein was assessed by using duplicated gel: the gel was stained for protein; photographed; and the density of corresponding lane minus background determined. (The whole blot and gel are shown in Additional file 6: Figure S5). (D) Nestin expression was quantified by using qRT-PCR in rat neural cells grown on ECM produced by SB623 transfected with either siTGM2 or siControl. Nestin levels on siControl-ECM were set on 1. The graph represents means from three experiments; error bar represents standard error of mean; *P < 0.05. (E) Rat nestin mRNA expression in cocultures with either siTGM2 or siControl transfectants. The graph represents mean from two experiments.