Journal: Stem Cell Research & Therapy

Article Title: Isolation and characterisation of AFP + /DLK1 + double-positive hepatic stem/progenitor-like cells from fibrotic partial hepatectomy model

doi: 10.1186/s13287-025-04728-1

Figure Lengend Snippet: Schematic presentation of the strategy for isolating AFP + /DLK1 + double-positive cells from the fibrotic PHx model

Article Snippet: Cells were then blocked with 2.5% BSA for 1 h. The cells were subsequently incubated with primary antibodies against AFP (1:200; Proteintech-cn, Wuhan, 14550-1-AP) and DLK1 (1:200; MBL, Beijing, D187-3) at 4 °C overnight, followed by incubation with secondary antibodies for 1 h at room temperature in the dark (1:1000, Abcam, USA).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Isolation and characterisation of AFP + /DLK1 + double-positive hepatic stem/progenitor-like cells from fibrotic partial hepatectomy model

doi: 10.1186/s13287-025-04728-1

Figure Lengend Snippet: Pre-injection of CCl 4 promotes liver regeneration after 2/3 PHx. a Schematic drawing of mouse liver anatomy and positioning of silk threads for knots. b Liver /Body Weight Ratio (%) after PHx. n = 2 per group. c Level of serum ALT and AST after PHx. n = 3 per group. d Relative mRNA expression levels of Afp , Dlk1 , Lgr5 , Ki67 , HNF4α , Alb , Krt-8 , Krt-18 , Krt7 ,and Krt-19 in regenerating livers after PHx. Values are normalised to Actb. n = 3 per group. e AFP (green), DLK1 (red), and DAPI (blue) immunofluorescence analyses in regenerating livers after PHx on the fibrotic model. Scale bars: 100 μm. f Relative fluorescence intensity (%). Statistical analysis was undertaken with Two-way ANOVA; (* p < 0.05; ** p < 0.01; *** p < 0.001, **** p < 0.0001.)

Article Snippet: Cells were then blocked with 2.5% BSA for 1 h. The cells were subsequently incubated with primary antibodies against AFP (1:200; Proteintech-cn, Wuhan, 14550-1-AP) and DLK1 (1:200; MBL, Beijing, D187-3) at 4 °C overnight, followed by incubation with secondary antibodies for 1 h at room temperature in the dark (1:1000, Abcam, USA).

Techniques: Injection, Expressing, Immunofluorescence, Fluorescence

Journal: Stem Cell Research & Therapy

Article Title: Isolation and characterisation of AFP + /DLK1 + double-positive hepatic stem/progenitor-like cells from fibrotic partial hepatectomy model

doi: 10.1186/s13287-025-04728-1

Figure Lengend Snippet: Isolation of AFP + /DLK1 + Double-Positive Cells from regenerating liver by density gradient differential centrifugation. a Program diagram of density gradient differential centrifugation. Whole liver cells obtained by enzymatic digestion (as shown in ①). By lowspeed centrifugation (50× g , 5 min), liver parenchymal cells are deposited at the bottom of the centrifuge tube, and liver nonparenchymal cells are suspended in the supernatant (as shown in ②). The supernatant is collected and centrifuged at medium speed (200× g , 5 min) to obtain nonparenchymal cell masses (as shown in ③). The nonparenchymal cells are placed in the top layer of the gradient centrifuge solution (as shown in ④). By differential centrifugation (600× g , 10 min, sped up by 9, and sped down by 1), all kinds of cells are separated into different Percoll layers (as shown in ⑤). Cells are collected from 50%–70% Percoll layer and HSPClike cells obtained by centrifugation (250× g , 5 min) (as shown in ⑥). b Flow cytometry analysis of AFP + /DLK1 + double-positive cells in each Percoll layer. c Percentage of AFP + /DLK1 + double-positive cells (by flow cytometry). d Relative mRNA expression levels of Afp , Dlk1 , Lgr5 , and Ki67 in each Percoll layer cells. Values are normalised to Actb. n = 3 per group. e AFP (green), DLK1 (red), and DAPI (blue) immunofluorescence analyses in the 50%– 70% layer. Scale bars:20 μm. f Relative fluorescence intensity (%). Statistical analysis was undertaken with Two-way ANOVA; (* p < 0.05; **** p < 0.0001.)

Article Snippet: Cells were then blocked with 2.5% BSA for 1 h. The cells were subsequently incubated with primary antibodies against AFP (1:200; Proteintech-cn, Wuhan, 14550-1-AP) and DLK1 (1:200; MBL, Beijing, D187-3) at 4 °C overnight, followed by incubation with secondary antibodies for 1 h at room temperature in the dark (1:1000, Abcam, USA).

Techniques: Isolation, Centrifugation, Gradient Centrifugation, Flow Cytometry, Expressing, Immunofluorescence, Fluorescence