Immunofluorescence was performed on cells grown directly on slides as previously described (Chadwick and Willard 2002 (link)). Except, the maximum slide seed area was delimited using a Super PAP Pen (IM3580, Beckman Coulter®) and coated with 700µl of Matrigel™ or 0.1% Gelatin prior to seeding hESCs or EBOGs, respectively. Additionally, hESCs were allowed to adhere for 1hr in complete StemPro® media supplemented with ROCK Inhibitor Y-27632 (ROCKi, SCM075, Millipore™), the media was then replaced with fresh complete StemPro® media (-ROCKi), and allowed to recover overnight. Antibodies used for indirect Immunofluorescence included rabbit anti-H3K4me2 (07–030, EMD Millipore), mouse anti-H3K27me3 (ab6002, abcam), rabbit anti-NANOG (3580S, Cell Signaling Technology), mouse anti-SSEA4 (4755S, Cell Signaling Technology), goat anti-SOX2 (AF2018, R&D Systems), and rabbit anti-OCT4 (2750S, Cell Signaling Technology). Conjugate secondary antibodies (Alexa-Fluor®) were obtained from Life Technologies Corporation. DNA was counterstained using the VECTASHIELD® mounting medium with DAPI from VECTOR Laboratories. Imaging was performed on an Olympus IX71 operated by the DeltaVision pDV, deconvolved with softWoRx 5.5.1 (DeltaVision), and compiled using Adobe Photoshop CS6 (Adobe Systems).
Goat anti sox2 af2018
Manufactured by R&D Systems
Goat anti-SOX2 (AF2018) is an antibody product developed by R&D Systems. The antibody is specific for the SOX2 (SRY-Box Transcription Factor 2) protein.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti h3k4me2, by Merck Group (1 mentions)
Rabbit anti oct4, by Cell Signaling Technology (1 mentions)
Mouse anti h3k27me3, by Abcam (1 mentions)
Mouse anti ssea4, by Cell Signaling Technology (1 mentions)
Rabbit anti nanog, by Cell Signaling Technology (1 mentions)
Vectashield mounting medium, by Vector Laboratories (1 mentions)
Hoechst 33342, by Thermo Fisher Scientific (1 mentions)
Mouse anti gfp, by Merck Group (1 mentions)
A21202, by Thermo Fisher Scientific (1 mentions)
A21203, by Thermo Fisher Scientific (1 mentions)
A11058, by Thermo Fisher Scientific (1 mentions)
Mouse anti mcherry, by Takara Bio (1 mentions)
2 protocols using goat anti sox2 af2018
1
Immunofluorescence Staining of Stem Cells
2
Immunohistochemistry for mCherry, Sox2, and GFP
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Immunohistochemistry was performed as described previously17 (link). In brief, sections on glass slides were treated in 10 mM Tris–HCl pH 9.0, 1 mM EDTA at 95 °C for one minute, incubated with the blocking buffer (10% normal donkey serum, 0.1% Triton-X100 in PBS), incubated with the primary antibodies at 4 °C overnight, washed, incubated with the secondary antibodies at room temperature for 1 h, and washed. The following antibodies were used; mouse anti-mCherry (#632543, Clontech), goat anti-Sox2 (AF2018, R&D Systems), mouse anti-GFP (#11814460001, Sigma), anti-mouse IgG conjugated to Alexa 594 (A21203, Molecular Probe) and anti-mouse IgG conjugated to Alexa 488 (A21202, Molecular Probe), anti-goat IgG conjugated to Alexa 594 (A11058, Molecular Probe). Hoechst 33342 (H1399, Molecular Probe) was used for nuclear staining.
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