box protein 6 pax6  (Developmental Studies Hybridoma Bank)

Journal: Stem Cell Research & Therapy

Article Title: Grafted c-kit + /SSEA1 − eye-wall progenitor cells delay retinal degeneration in mice by regulating neural plasticity and forming new graft-to-host synapses

doi: 10.1186/s13287-016-0451-8

Figure Lengend Snippet: Progenitor characteristics of mouse eye-wall c-kit + /SSEA1 − cells. ( A ) Representative flow cytometry plots showing the percentage of c-kit-positive SSEA1-negative cells. Gating was established based on cells stained with isotype-matched APC and FITC antibodies ( ISO ; left panel ). Representative flow cytometry plots showed that c-kit + /SSEA1 − cells represented approximately 0.82% of the total population ( right panel ). ( B ) Phase-contrast image of representative c-kit + /SSEA1 − cells in culture. ( C ) Representative image of immunofluorescence staining for c-kit + ( green ) with 4′,6-diamidino-2-phenylindole ( DAPI ; blue ). ( D–H ) Representative images of immunofluorescence for RPC markers ( red ) and DAPI ( blue ), showing that cells express Nestin ( D ), retina homeobox protein Rx ( Rax ; E ), SRY-box 2 ( Sox2 ; F ), Orthodenticle Homeobox 2 ( Otx2 ; G ), and paired box protein 6 ( Pax6 ; H ). ( D′–H′ ) Representative flow cytometry plots showing expression in the FITC channel for Nestin (74%; D′ ), Rax (98.8%; E′ ), Sox2 (97.7%; F′ ), Otx2 (99.8%; G′ ), and Pax6 (95.7%; H′ ). Scale bars represent 50 μm (Color figure online)

Article Snippet: The primary antibodies used were as follows: anti-c-kit at 1:200 (Cell Signaling Technology, Danvers, MA, USA), anti-nestin at 1:200 (Abcam, Cambridge, MA, USA), anti-retina homeobox protein Rx (Rax) at 1:200 (Abcam), anti-SRY (sex determining region Y)-box 2 (Sox2) at 1:500 (Abcam), anti-orthodenticle homeobox 2 (Otx2) at 1:400 (Abcam), anti-paired box protein 6 (Pax6) at 1:200 (Abcam), anti-Ki67 at 1:250 (Abcam), anti-telomerase reverse transcriptase (TERT) at 1:200 (EMD Millipore), anti-recoverin at 1:1000 (EMD Millipore), anti-rhodopsin at 1:1000 (Abcam), anti-protein kinase C alpha (PKCα) at 1:250 (Abcam), anti-calbindin at 1:200 (Abcam), anti-glutamate decarboxylase 65 & 67 (GAD) at 1:500 (Abcam), anti-choline acetyltransferase (ChAT) at 1:100 (Abcam), anti-glutamine synthetase (GS) at 1:250 (Abcam), anti-glial fibrillary acidic protein (GFAP) at 1:100 (Abcam), anti-microphthalmia-associated transcription factor (MITF) at 1:100 (Abcam), anti-calponin at 1:100 (Abcam), anti-von Willebrand factor (vWF) at 1:100 (EMD Millipore), anti-synaptophysin at 1:100 (EMD Millipore), and anti-postsynaptic density-95 (PSD-95) at 1:100 (EMD Millipore).

Techniques: Flow Cytometry, Staining, Immunofluorescence, Expressing

Journal: Stem Cell Research & Therapy

Article Title: Grafted c-kit + /SSEA1 − eye-wall progenitor cells delay retinal degeneration in mice by regulating neural plasticity and forming new graft-to-host synapses

doi: 10.1186/s13287-016-0451-8

Figure Lengend Snippet: Transdifferentiation capability of eye-wall c-kit + /SSEA1 − progenitor cells. ( A ) Day 1 in which the medium of c-kit + /SSEA1 − cells was switched to the RPE differentiation medium. ( B ) Pigment ( arrowhead ) appeared after 4–8 weeks. ( C ) Pigment ( arrow ) could be seen in the dish. Representative immunostaining images showing cells positive for paired box protein 6 ( Pax6 ; D ), microphthalmia-associated transcription factor ( MITF ; E ), Calponin ( F ), and von Willebrand factor ( vWF ; G ). Differentiated cells were stained for markers shown in the FITC and APC channels. Representative flow cytometry plots showing the percentages of cells positive for Pax6 (26.6%; D′ ), MITF (16.9%; E′ ), Calponin (27.3%; F′ ), and vWF (25.6%; G′ ). DAPI 4′,6-diamidino-2-phenylindole. Scale bars represent 50 μm for all the images (Color figure online)

Article Snippet: The primary antibodies used were as follows: anti-c-kit at 1:200 (Cell Signaling Technology, Danvers, MA, USA), anti-nestin at 1:200 (Abcam, Cambridge, MA, USA), anti-retina homeobox protein Rx (Rax) at 1:200 (Abcam), anti-SRY (sex determining region Y)-box 2 (Sox2) at 1:500 (Abcam), anti-orthodenticle homeobox 2 (Otx2) at 1:400 (Abcam), anti-paired box protein 6 (Pax6) at 1:200 (Abcam), anti-Ki67 at 1:250 (Abcam), anti-telomerase reverse transcriptase (TERT) at 1:200 (EMD Millipore), anti-recoverin at 1:1000 (EMD Millipore), anti-rhodopsin at 1:1000 (Abcam), anti-protein kinase C alpha (PKCα) at 1:250 (Abcam), anti-calbindin at 1:200 (Abcam), anti-glutamate decarboxylase 65 & 67 (GAD) at 1:500 (Abcam), anti-choline acetyltransferase (ChAT) at 1:100 (Abcam), anti-glutamine synthetase (GS) at 1:250 (Abcam), anti-glial fibrillary acidic protein (GFAP) at 1:100 (Abcam), anti-microphthalmia-associated transcription factor (MITF) at 1:100 (Abcam), anti-calponin at 1:100 (Abcam), anti-von Willebrand factor (vWF) at 1:100 (EMD Millipore), anti-synaptophysin at 1:100 (EMD Millipore), and anti-postsynaptic density-95 (PSD-95) at 1:100 (EMD Millipore).

Techniques: Immunostaining, Staining, Flow Cytometry