Journal: Cancer drug resistance (Alhambra, Calif.)

Article Title: The uniqueness of ABCB5 as a full transporter ABCB5FL and a half-transporter-like ABCB5β.

doi: 10.20517/cdr.2024.56

Figure Lengend Snippet: Figure 2. Tissue and intracellular localization of ABCB5FL and ABCB5β. (A) Representation of human ABCB5 tissue localization based on ABCB5FL and ABCB5β mRNA detection from

Article Snippet: In melanoma or retinoblastoma cell culture, ABCB5 protein (600-401-A77 antibody from Rockland) or mRNA expression did not correlate with CD271 or CD133 expression[48,56].

Techniques:

Journal: Nature

Article Title: ABCB5 is a limbal stem cell gene required for corneal development and repair

doi: 10.1038/nature13426

Figure Lengend Snippet: a, Cornea and LSC niche. b, BrdU detection (8-week chase). c, d, Immunofluorescence (×60 magnification) (c) and flow cytometric staining (gating based on control-staining) (d) for Abcb5/BrdU co-expression in mouse limbus. DAPI, 4′,6-diamidino-2-phenylindole. Abcb5 and BrdU co-expression data in d represent analyses of n = 4 mice per group (mean ± standard error of the mean (s.e.m.)). The experiment was performed three times. Data were analysed using the unpaired t-test, P < 0.001. e, ABCB5 positivity in human limbus (palisades of Vogt, ×20 magnification), with negativity in central cornea. f, ABCB5/p63α co-expression in human limbus. Quantitative analysis of ABCB5 monoclonal antibody and ΔNp63α/TAp63α epitope-binding antibody co-expression was performed using limbal epithelial cells from n = 4 eyes. The experiment was performed twice. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m., P < 0.05. The quantitative analysis of ABCB5 monoclonal antibody and ΔNp63α,β,γ epitope binding antibody co-expression was performed using limbal epithelial cells from n = 4 eyes. The experiment was performed twice. Data were analysed using the Mann–Whitney test. Data are shown as mean ± s.e.m., P < 0.05. g, ABCB5/KRT12 co-expression. FSC, forward scatter. h, ABCB5 in LSCD patients (×20 magnification). Bar graph shows per cent ABCB5+ cells in control donors versus LSCD patients. Quantitative analysis of ABCB5 expression in n = 2 control and n = 2 LSCD specimens was performed using n = 8 sections per patient/control. All epithelial cells were counted in each section. A total of 2,031 and 2,051 epithelial cells were counted in patient 1 and donor 1, respectively. A total of 563 and 2,662 epithelial cells were counted in patient 2 and donor 2, respectively. Data were analysed using the unpaired t-test. Error bars indicate s.e.m. *P < 0.05, ***P < 0.001.

Article Snippet: Abcb5 wild-type and Abcb5 knockout cell lysates were immunoblotted using monoclonal ABCB5 antibody 3C2-1D12 (ref. 6 ) (5.5 μg ml −1 ), a rabbit polyclonal N-terminus-targeted ABCB5 antibody (1:100 dilution) (Abgent), or an α-tubulin rabbit polyclonal antibody (1:5,000 dilution) (Abcam).

Techniques: Immunofluorescence, Staining, Expressing, Binding Assay, MANN-WHITNEY

Journal: Nature

Article Title: ABCB5 is a limbal stem cell gene required for corneal development and repair

doi: 10.1038/nature13426

Figure Lengend Snippet: a, Abcb5 locus and protein topology (transmembrane protein topology with a hidden Markov model (TMHMM), knockout deletion in red). mAb, monoclonal antibody. b, Abcb5 knockout strategy. BAC, bacterial artificial chromosome. c, PCR and western blot in wild-type (WT) versus knockout (KO) mice. HT, heterozygous. d, e, Abcb5 immunofluorescence staining (d) and in situ hybridization (e) of wild-type and knockout limbus and cornea (×20 magnification). f–h, Slit lamp and haematoxylin and eosin (H&E; ×40 magnification) (f), cellularity (g) and LC-biotin diffusion and protein expression (h) analyses. i, Analysis of wild-type and knockout corneas following debridement wounding. j, k, Ki67 immunofluorescence (j) and TdT-mediated dUTP nick end labelling (TUNEL) (k) staining. g–k, ×20 magnification. g, The numbers of viable epithelial cells in Abcb5 knockout versus Abcb5 wild-type murine central cornea were derived from the analysis of n = 8 mice per group (left bar graph). The experiment was performed four times. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m., P < 0.05. In the right bar graph, the numbers of viable epithelial cells in Abcb5 knockout versus Abcb5 wild-type murine limbus were derived from the analysis of n = 6 mice per group. The experiment was performed three times. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m., P = not significant. h, Quantitative analyses of Pax6 expression in limbus and cornea of Abcb5 knockout versus Abcb5 wild-type mice were performed using n = 3 mice per group. The experiment was performed three times. Ten thousand cells per experiment were counted for each group. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m., P < 0.05. Quantitative analyses of Krt12 expression in cornea were performed using n = 2 mice per group. The experiment was performed twice. Ten thousand cells per experiment were counted for each group. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m., P < 0.05. i, The numbers of DAPI+ cells per section in Abcb5 wild-type versus Abcb5 knockout mice (mean ± s.e.m.) were derived from n = 6 mice per group. The experiment was performed twice. Within the standardized area (shown in Extended Data Fig. 4f), all corneal epithelial cells were counted in at least n = 3 consecutive composite cross-sections. Data were analysed using the unpaired t-test, P < 0.001. j, The percentages of Ki67+ epithelial cells in limbus and cornea of Abcb5 knockout versus Abcb5 wild-type mice (mean ± s.e.m.) were determined using n = 4 mice per group. The experiment was performed twice. Within a standardized area, all limbal epithelial cells were counted in at least n = 3 consecutive cross-sections. Data were analysed using the unpaired t-test, P < 0.001 for limbus and P < 0.05 for cornea. k, The percentages of TUNEL+ epithelial cells in limbus or cornea of Abcb5 knockout versus Abcb5 wild-type mice (mean ± s.e.m.) were determined using n = 4 mice per group. The experiment was performed twice. Within a standardized area, all limbal epithelial cells were counted in at least n = 3 consecutive cross-sections. Data were analysed using the unpaired t-test, P < 0.001 for limbus and P < 0.001 for cornea. Error bars indicate s.e.m. *P < 0.05, ***P < 0.001. NS, not significant.

Article Snippet: Abcb5 wild-type and Abcb5 knockout cell lysates were immunoblotted using monoclonal ABCB5 antibody 3C2-1D12 (ref. 6 ) (5.5 μg ml −1 ), a rabbit polyclonal N-terminus-targeted ABCB5 antibody (1:100 dilution) (Abgent), or an α-tubulin rabbit polyclonal antibody (1:5,000 dilution) (Abcam).

Techniques: Knock-Out, Western Blot, Immunofluorescence, Staining, In Situ Hybridization, Diffusion-based Assay, Expressing, TUNEL Assay, Derivative Assay

Journal: Nature

Article Title: ABCB5 is a limbal stem cell gene required for corneal development and repair

doi: 10.1038/nature13426

Figure Lengend Snippet: a, b, Murine syngeneic grafts (a) and 5-week (b) or 13-month (c) human xenografts to LSCD-induced mice. Grafts contained either no donor cells (row 2), ABCB5− cells (row 3), unsegregated cells (row 4), or ABCB5+ cells (row 5). a–c, Untreated C57BL/6 (a) and NSG (b, c) corneas (without LSCD) are shown in row 1. b, Bottom right, RT–PCR detection of human donor cells. d, Human KRT12+ cells (red) in recipient corneas of ABCB5+ human grafts at 13 months. Magnification: columns 2 and 4: ×20; column 3: ×40. Scale bars: 100 μm. The transplantation experiments shown in a and b were performed in n = 5 mice per group. The experiment was performed twice. For KRT12 expression analyses, all corneal epithelial cells within a standardized area (Extended Data Fig. 8b) were counted in at least n = 3 consecutive cross-sections from n = 4 replicate mice per group. Data were analysed using the one-way ANOVA and Bonferroni multiple comparisons tests. Data are shown as mean ± s.e.m. c, The transplantation experiments were performed in n = 5 mice per group. The experiment was performed twice. For KRT12 expression analyses, all corneal epithelial cells within a standardized area (Extended Data Fig. 9b) were counted in at least n = 3 consecutive cross-sections from n = 4 replicate mice per group. Data were analysed using the one-way ANOVA and Bonferroni multiple comparisons tests. Data are shown as mean ± s.e.m. d, Human KRT12 expression was analysed in n = 4 mice per group. Within a standardized area (Extended Data Fig. 9b), all corneal epithelial cells were counted in at least n = 3 consecutive cross-sections. Data were analysed using the unpaired t-test. Data are shown as mean ± s.e.m. Error bars indicate s.e.m. **P < 0.01, ***P < 0.001. ND, not detected; NS, not significant.

Article Snippet: Abcb5 wild-type and Abcb5 knockout cell lysates were immunoblotted using monoclonal ABCB5 antibody 3C2-1D12 (ref. 6 ) (5.5 μg ml −1 ), a rabbit polyclonal N-terminus-targeted ABCB5 antibody (1:100 dilution) (Abgent), or an α-tubulin rabbit polyclonal antibody (1:5,000 dilution) (Abcam).

Techniques: Reverse Transcription Polymerase Chain Reaction, Transplantation Assay, Expressing