rabbit anti ck7  (Proteintech)

Journal: bioRxiv

Article Title: The Hippo/YAP Pathway Mediates the De-differentiation of Corneal Epithelial Cells into Functional Limbal Epithelial Stem Cells In Vivo

doi: 10.1101/2024.06.11.596348

Figure Lengend Snippet: ( A , C ) The whole-mount F-actin staining of normal cornea and cornea at 12h after the removal of limbal epithelium. The cellular area of peripheral corneal epithelial cells was quantified. Data are the mean ± SD, n=25 cells; statistical analysis are performed by Student’s unpaired t-test, and p value is shown. ( B , D ) The EdU-positive proliferative epithelial cells at limbus (Lim.), peripheral cornea (Per.) and central cornea (Cen.) at indicated time points after the removal of limbal epithelium were quantified. Data are the mean ± SD, n=8 zones. ( E ) The ocular surface between the conjunctiva and central cornea was regionalized into four parts, namely 1, 2, 3, and 4 as shown. The different-sized zones of limbal and corneal epithelium were scratched off, resulting in different outcomes of the competition between corneal and conjunctival epithelial cells for the limbus. The whole-mount CK12 or CK7 immunostaining was performed against corneas with different-sized wounds. Dashed circles indicate the limbus. Scale bars, 20um ( A ) and 100um ( B ).

Article Snippet: The following primary antibodies were used: rabbit anti-CK12 monoclonal antibody (Abcam, ab185627; 1:400), rabbit anti-CK7 monoclonal antibody (Abcam, ab181598; 1:400), rabbit anti-connexin 43 (Cx43) monoclonal antibody (CST, #3512; 1:80), rabbit anti-ApoE monoclonal antibody (Abcam, ab183596; 1:400), rabbit anti-CK14 monoclonal antibody (Abcam, ab119695; 1:200), rabbit anti-deltaN-p63 polyclonal antibody (BioLegend, 619002; 1:400), rabbit anti-p75NTR monoclonal antibody (CST, #8238; 1:800), rabbit anti-CD63 polyclonal antibody (Bioworld, BS72936; 1:400), rabbit anti-TSPAN7 polyclonal antibody (Proteintech, 18695-1-AP; 1:100), rabbit anti-IFITM3 monoclonal antibody (CST, #59212; 1:100), mouse anti-ATF3 monoclonal antibody (Santa Cruz, sc-518032; 1:100), rabbit anti-YAP monoclonal antibody (CST, #14074; 1:100), rabbit anti-p-YAP-Ser127 monoclonal antibody (Abcam, ab76252; 1:100), rabbit anti-active-YAP (non-phosphorylated YAP) monoclonal antibody (Abcam, ab205270; 1:200), rabbit anti-Ki67 monoclonal antibody (CST, #9129; 1:100), rabbit anti-p-Histone H3-Ser10 (pH3) monoclonal antibody (CST, #53348; 1:100) and mouse anti-CK15 monoclonal antibody (Santa Cruz, sc-47697; 1:200).

Techniques: Staining, Immunostaining

Journal: bioRxiv

Article Title: The Hippo/YAP Pathway Mediates the De-differentiation of Corneal Epithelial Cells into Functional Limbal Epithelial Stem Cells In Vivo

doi: 10.1101/2024.06.11.596348

Figure Lengend Snippet: ( A ) Experimental strategy to resolve the regeneration of LESCs after innate stem cells ablation in vivo . The transprarency, conjunctivalization and two consecutive central corneal wound healing are shown in . The expressions of LESCs markers and dynamics of stripes/clones are shown in and - , respectively. The different-sized wounds (small, medium and large) of central corneal epithelium in mice are also shown. ( B ) The limbal epithelium and marginal corneal and conjunctival epithelium of 6-month-old C56BL/6J mice were removed by surgery. Fluorescein sodium staining and bright-field images of injured eyes at indicated days are shown. Whole-mount normal and LESCs-ablation (LESCs-Ab; 6 months) corneas were immunostained for conjunctival epithelial marker CK7 and corneal epithelial marker CK12. ( C ) Experimental strategy of two consecutive central corneal wound healing of normal and LESCs-ablation corneas. ( D ) Fluorescein sodium images and whole-mount staining of normal and LESCs-ablation corneas for EdU (cell proliferation) and CK12 after the first and second wound healing. Dashed circles indicate the limbus, which is localized based on the location of iris and ciliary body in the bright field. These EdU-positive proliferative cells at 8 zones of peripheral cornea are quantified. Data are the mean ± SD; statistical analysis are performed by Student’s unpaired t-test, and p values are shown.

Article Snippet: The following primary antibodies were used: rabbit anti-CK12 monoclonal antibody (Abcam, ab185627; 1:400), rabbit anti-CK7 monoclonal antibody (Abcam, ab181598; 1:400), rabbit anti-connexin 43 (Cx43) monoclonal antibody (CST, #3512; 1:80), rabbit anti-ApoE monoclonal antibody (Abcam, ab183596; 1:400), rabbit anti-CK14 monoclonal antibody (Abcam, ab119695; 1:200), rabbit anti-deltaN-p63 polyclonal antibody (BioLegend, 619002; 1:400), rabbit anti-p75NTR monoclonal antibody (CST, #8238; 1:800), rabbit anti-CD63 polyclonal antibody (Bioworld, BS72936; 1:400), rabbit anti-TSPAN7 polyclonal antibody (Proteintech, 18695-1-AP; 1:100), rabbit anti-IFITM3 monoclonal antibody (CST, #59212; 1:100), mouse anti-ATF3 monoclonal antibody (Santa Cruz, sc-518032; 1:100), rabbit anti-YAP monoclonal antibody (CST, #14074; 1:100), rabbit anti-p-YAP-Ser127 monoclonal antibody (Abcam, ab76252; 1:100), rabbit anti-active-YAP (non-phosphorylated YAP) monoclonal antibody (Abcam, ab205270; 1:200), rabbit anti-Ki67 monoclonal antibody (CST, #9129; 1:100), rabbit anti-p-Histone H3-Ser10 (pH3) monoclonal antibody (CST, #53348; 1:100) and mouse anti-CK15 monoclonal antibody (Santa Cruz, sc-47697; 1:200).

Techniques: In Vivo, Clone Assay, Staining, Marker

Journal: bioRxiv

Article Title: The Hippo/YAP Pathway Mediates the De-differentiation of Corneal Epithelial Cells into Functional Limbal Epithelial Stem Cells In Vivo

doi: 10.1101/2024.06.11.596348

Figure Lengend Snippet: ( A ) Experimental strategy of the localized NaOH application to destroy the limbal niche, which allows the examination of the fate of CECs either with or without an intact limbal niche on the same cornea. ( B ) The whole-mount CK7 immunostaining against corneas of scratched limbus either with or without NaOH application. Dashed circles indicate the limbus. ( C ) The frozen-section CK7 and CK12 immunostaining against cornea of scratched limbus either with or without NaOH application at 10 and 20 days. ( D, E ) The frozen-section Cx43 and ApoE immunostaining against cornea of scratched limbus either with or without NaOH application at 6 and 10 days. The percentage of ApoE + LESCs and the mean fluorescence intensity (MFI) of Cx43 were quantified. Data are the mean ± SD, n=5 biological replicates; statistical analysis were performed by paired one-way ANOVA with Tukey’s test. Scale bars, 20 um.

Article Snippet: The following primary antibodies were used: rabbit anti-CK12 monoclonal antibody (Abcam, ab185627; 1:400), rabbit anti-CK7 monoclonal antibody (Abcam, ab181598; 1:400), rabbit anti-connexin 43 (Cx43) monoclonal antibody (CST, #3512; 1:80), rabbit anti-ApoE monoclonal antibody (Abcam, ab183596; 1:400), rabbit anti-CK14 monoclonal antibody (Abcam, ab119695; 1:200), rabbit anti-deltaN-p63 polyclonal antibody (BioLegend, 619002; 1:400), rabbit anti-p75NTR monoclonal antibody (CST, #8238; 1:800), rabbit anti-CD63 polyclonal antibody (Bioworld, BS72936; 1:400), rabbit anti-TSPAN7 polyclonal antibody (Proteintech, 18695-1-AP; 1:100), rabbit anti-IFITM3 monoclonal antibody (CST, #59212; 1:100), mouse anti-ATF3 monoclonal antibody (Santa Cruz, sc-518032; 1:100), rabbit anti-YAP monoclonal antibody (CST, #14074; 1:100), rabbit anti-p-YAP-Ser127 monoclonal antibody (Abcam, ab76252; 1:100), rabbit anti-active-YAP (non-phosphorylated YAP) monoclonal antibody (Abcam, ab205270; 1:200), rabbit anti-Ki67 monoclonal antibody (CST, #9129; 1:100), rabbit anti-p-Histone H3-Ser10 (pH3) monoclonal antibody (CST, #53348; 1:100) and mouse anti-CK15 monoclonal antibody (Santa Cruz, sc-47697; 1:200).

Techniques: Immunostaining, Fluorescence