Journal: bioRxiv

Article Title: CD133 + Intercellsome Mediates Direct Cell-Cell Communication to Offset Intracellular Signal Deficit

doi: 10.1101/2022.05.16.492226

Figure Lengend Snippet: a , General morphology of livers 2 days after PHx. b , H&E staining of liver sections 2 days after PHx. c , d , Ki67 + percentages in HNF4α + hepatocytes ( c ) and HNF4α − NPCs ( d ) 2 days after PHx. Means ± SEM are shown, n=3 per group, **P < 0.01 (two-tailed unpaired t test). n.s., not significant. e , EpCAM was highly expressed in biliary epithelial cells (arrow), but not in the colony (dashed line). Stellate cells (GFAP) were not altered around the colony. f , CK19 was highly expressed in biliary epithelial cells (arrow), but not in the colony (dashed line). CD133 was highly expressed in the colony. g , Sox9 and CD44 were highly expressed in biliary epithelial cells (arrowhead), but was not upregulated in the colony (dashed line) compared to surrounding hepatocytes. h , AFP showed no difference between the colony (dashed line) and the surrounding tissue. i , Liver/body weight ratios after PHx. Means ± SEM from 3 or more mice analyzed for each time point and group are shown. j , k , Immunofluorescent staining of SKO liver sections 3 weeks after PHx. Colonies were at various sizes and locations as shown by arrows in j . Macroscopic colonies were found as shown by arrows and dashed lines in k . Immunofluorescent image in k corresponds to the magenta arrow in the liver image. PV, portal vein; CV, central vein. l , Vasculature shown by PECAM did not distinguish the colony (arrow). m , HGF expressed by NPCs was not concentrated in the colony (dashed line). Scale bars, 1 cm ( a ) 100 μm ( b , e - g , i - l ).

Article Snippet: Antibodies for Ki67 (14-5698-80; eBioscience), HNF4α (sc-8987; Santa Cruz Biotechnology), mouse CD133 (14-1331-82; eBioscience), human CD133 (86781; CST), E-cadherin (sc-7870; Santa Cruz Biotechnology), GFP (04404-84; Nacalai Tesque), Shp2 (sc-280; Santa Cruz Biotechnology), Porcupine (ab105543; abcam), β-catenin (sc-7199; Santa Cruz Biotechnology), CHMP2B (ab157208; abcam), HuR (ab200342; abcam, and 66549-1-Ig; proteintech), VE-Cad (AF1002; R&D SYSTEMS), BrdU (555627; BD Biosciences), PECAM (13-0311-81; eBioscience), HGF (ab83760; abcam), GFAP (Z0334; Dako), EpCAM (552370; BD Biosciences), Sox9 (AB5535: Millipore), CD44v6 (GTX75661; GeneTex) and AFP (AF5369; R&D SYSTEMS) were used as primary antibodies.

Techniques: Staining, Two Tailed Test

Journal: EBioMedicine

Article Title: Tumour-specific activation of a tumour-blood transport improves the diagnostic accuracy of blood tumour markers in mice.

doi: 10.1016/j.ebiom.2024.105178

Figure Lengend Snippet: Fig. 1: Intravenously injected iRGD increases blood AFP levels in HCC-bearing mice. (a) Experimental setup to study the effect of iRGD on blood AFP levels in Huh-7 xenografted nude mice. (b and c) Blood AFP levels before and after intravenous injection of iRGD (b, c, n = 12), RGD control peptide (c (n = 6)), and PBS (c (n = 6)) in mice with HepG2 xenografts. Human AFP was not detectable in the blood of mice without tumours. In (c) the fold increase of AFP with pre-injection level set to 1; lines and error bars represent geometric means and 95% CI. (d) Experimental setup to study the effect of iRGD on blood AFP levels in TGFα/c-myc HCC mice. (e and f) iRGD specifically increased the blood AFP levels in TGFα/c-myc HCC mice. TGFα/c-myc mice (20–24 weeks old) with HCC according to MRI or without HCC (f) were intravenously injected with iRGD (e, f), RGD control peptide (f) or PBS (f). Data are fold changes of blood AFP due to the treatments (n = 48: iRGD; RGD control peptide: n = 34; PBS: n = 15); lines and error bars indicate medians and 95% CI. Dashed line: upper 95% CI increase in blood AFP in the PBS-injected HCC mice. (g) AFP expression in HCCs of TGFα/c-myc mice. HCCs and liver tissues were excised from TGFα/c-myc mice. Pairs of the tissue lysates were analysed for AFP and β-actin content by immunoblotting. Band densities were measured densitometrically. The ratio of AFP/β-actin in the livers was set to 1. (h) iRGD-induced accumulation of Evans blue in HCCs in TGFα/c-myc mice with HCC. TGFα/c-myc mice with HCCs were co- injected with iRGD or PBS (n = 12 per group) and Evans blue (EB). The dye content of the tumours was related to that in the livers; lines and error bars indicate geometric means (b, c, and h) or medians (e and f) and 95% CI; dashed line: upper 95% CI of the measured EB content of a HCC from the PBS-injected animals. (b): paired t test for log-transformed data; (c) One-way ANOVA with multiple comparison post-hoc test for log-transformed data; (e): Wilcoxon signed-rank test for log-transformed data; (f) Kruskal–Wallis test with Dunn’s multiple comparison post- hoc test; (h): two-sample t test. The indicated fold increase in (b, e and h) is the ratio of the geometric means with 95% CI.

Article Snippet: Immunoblotting of lysates obtained from pairs of liver and HCC tissue from TGFα/c-myc mice was performed as described previously.38 Gel-resolved proteins were electrotransferred to nitrocellulose membranes and incubated with antibodies raised against mouse-AFP (#AF5369, R&D Systems, Minneapolis, MN, RRID:AB_2258018) and anti-β-actin (#A2066, Sigma– Aldrich/Merck, RRID:AB_476693).

Techniques: Injection, Control, Expressing, Western Blot, Transformation Assay, Comparison

Journal: EBioMedicine

Article Title: Tumour-specific activation of a tumour-blood transport improves the diagnostic accuracy of blood tumour markers in mice.

doi: 10.1016/j.ebiom.2024.105178

Figure Lengend Snippet: Fig. 3: iRGD-induced elevation of the blood AFP concentration depends on NRP-1 and the tumour blood concentration gradient of AFP. (a) Anti-NRP-1 prevented iRGD-induced increase in the blood AFP concentration. TGFα/c-myc tumour mice that displayed a robust iRGD-induced increase in blood AFP level one week earlier were injected with anti-NRP-1 and the effect of iRGD on blood AFP level was determined (n = 3 per group). Fold increase of AFP with pre-injection level at the first time point was set to 1. Lines and error bars represent medians and 95% CI. (b– e) iRGD-induced elevation of the blood AFP level correlated negatively with the pre-injection blood AFP level in TGFα/c-myc mice (b), DEN- CCl4-HCC mice (c), mice with Huh-7 (d) or HepG2 xenografts (e). Spearman correlation r (b and c) and Pearson correlation r (d and e) with 95% CI, two tailed p-values and the log–log regression lines. (f) iRGD increased the blood AFP levels in HepG2 xenografted nude mice and low basal AFP (<67 ng/ml, n = 36, left panel), but not in animals with high basal AFP (>67 ng/ml, n = 12, right panel). Lines and error bars indicate geometric means and 95% CI. [(g) iRGD increased the blood AFP levels in TGFα/c-myc mice with HCC and normal basal AFP (<67 ng/ml, n = 36, left panel), but not in mice with elevated basal AFP (>67 ng, n = 12, right panel). Lines and error bars represent medians (left) or geometric means (right) with 95% CI. Significance was calculated with one sample t test (a, left) and the unpaired t test (a, right), paired t test (f and g, right) and Wilcoxon matched-pairs signed-rank test (g, left). The indicated fold increase in (f) is the geometric mean ratio with 95% CI. The indicated fold increase in (g) is the median of the ratios with 95% CI.

Article Snippet: Immunoblotting of lysates obtained from pairs of liver and HCC tissue from TGFα/c-myc mice was performed as described previously.38 Gel-resolved proteins were electrotransferred to nitrocellulose membranes and incubated with antibodies raised against mouse-AFP (#AF5369, R&D Systems, Minneapolis, MN, RRID:AB_2258018) and anti-β-actin (#A2066, Sigma– Aldrich/Merck, RRID:AB_476693).

Techniques: Concentration Assay, Injection, Two Tailed Test

Journal: International Journal of Nanomedicine

Article Title: Characteristics of magnetic labeling on liver tumors with anti-alpha-fetoprotein-mediated Fe 3 O 4 magnetic nanoparticles

doi: 10.2147/IJN.S30061

Figure Lengend Snippet: Information of in-vivo tests and biopsy tests in this work

Article Snippet: The dextran then reacted with the anti-AFP through −CH=N− to covalently conjugate anti-AFP antibodies (EA502-Q1053; EastCoast Bio, North Berwick, ME, USA) onto MNPs.

Techniques: Staining, H&E Stain

Journal: International Journal of Nanomedicine

Article Title: Characteristics of magnetic labeling on liver tumors with anti-alpha-fetoprotein-mediated Fe 3 O 4 magnetic nanoparticles

doi: 10.2147/IJN.S30061

Figure Lengend Snippet: Characteristics of anti-CEA MF, anti-AFP MF, and MF without antibody coatings: ( A ) the hydrodynamic diameters, ( B ) saturation magnetism, ( C ) the stability of antibody-mediated MFs. Abbreviations: CEA, carcinoembryonic-antigen; MF, magnetic fluid; AFP, alphafetaprotein.

Article Snippet: The dextran then reacted with the anti-AFP through −CH=N− to covalently conjugate anti-AFP antibodies (EA502-Q1053; EastCoast Bio, North Berwick, ME, USA) onto MNPs.

Techniques:

Journal: Genetics and Molecular Research

Article Title: Characterization of yolk sac proteins of Bos indicus cattle embryos

doi: 10.4238/2012.november.14.1

Figure Lengend Snippet: Figure 2. Membranes after ECL showing the molecular weight (MW) and samples of the yolk sac of embryos 2 and 3 at an estimated gestational period from 30 to 40 days. Red arrows indicate the α-fetoprotein in the samples 2:03 with a molecular weight of 70 kDa.

Article Snippet: The secondary antibody was also diluted in TTBS and applied to the membrane for 1 h. Mouse anti-human α-fetoprotein monoclonal antibody (AFP; Serotec, MCA1863HT), mouse anti-human transferrin antibody (BD, 612124) and rabbit anti-human α-1-antitrypsin polyclonal antibody (A1AT; Abcan, AB922) were applied as primary antibodies, and anti-mouse- and anti-rabbitperoxidase conjugate as secondary antibody (GE, NIF825).

Techniques: Molecular Weight

Journal: Frontiers in Physiology

Article Title: Pathogenesis of Molar Hypomineralisation: Hypomineralised 6-Year Molars Contain Traces of Fetal Serum Albumin

doi: 10.3389/fphys.2020.00619

Figure Lengend Snippet: A molecular approach for timestamping exposures of serum albumin to dental enamel. (A) Schematic showing three hypothetical stages where albumin could become incorporated in enamel on a 6-year molar ( yellow spot ), as considered in this study. These albumin exposures are designated developmental (0–3 years), eruptive (6 years), or artefactual during subsequent extraction, as indicated. (B) Serum levels of serum albumin (ALB; dark blue ) and alpha-fetoprotein (AFP; cyan ) in healthy individuals aged from 3-months postconception through to 1-year old, as indicated. Data were collated and averaged from 9 population studies as outlined in Methods and . It can be seen that at birth ( dotted line ), ALB is about 600-fold more concentrated than AFP. (C) The adult/fetal isoform ratio for serum albumin ( black line ), as derived from (B) , rises quasi-linearly through to 6-months of age ( cream box ). Sensitivity limits for the AFP immunoblot assay reached the early-post-natal period ( cyan box ). Note the isoform ratio, which spans 6-orders of magnitude (cf. log scale), offers strong resolution of age in early infancy.

Article Snippet: Human albumin (Sigma), alpha-fetoprotein (Lee Biosolutions), recombinant alpha-fetoprotein and albumin (both tagged with glutathione S-transferase; from Abnova), and rabbit polyclonal antibodies to human albumin (PAB10220 from Abnova), alpha-fetoprotein peptide (PAB12795 from Abnova) and whole alpha-fetoprotein (H00000174-D01 from Abnova), were obtained commercially.

Techniques: Extraction, Derivative Assay, Cream, Western Blot

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