Zonae pellucidae were removed by a brief incubation with acidic Tyrode’s solution, and zona pellucida-free embryos were washed in phosphate-buffered saline (PBS), fixed in freshly prepared 2% paraformaldehyde for 20 min at room temperature, and washed twice in PBS. Afterward, the embryos were permeabilized in 0.5% Triton X-100 for 25 min, washed three times in PBS, and then blocked with 5% donkey serum for 1 h. Samples were incubated with SIN3A antibody (Cat #3479, Abcam), TRF2 antibody (Cat #ab108997, Abcam), or rabbit polyclonal anti-γH2AX antibody (Cat #80312, Cell Signaling Technology) diluted 1: 100 in blocking solution at 4°C overnight, washed three times, incubated with FITC donkey anti-mouse IgG or FITC anti-rabbit IgG for 1 h at room temperature in the dark, washed again, and finally mounted with VECTASHIELD mounting medium and exposed to 0.5 μg/mL 4, 6-diamidino-2-phenylindole (DAPI, 1 μg/mL, Thermo Fisher Scientific). Images were captured with a ZEISS microscope equipped with fluorescence (Celldiscoverer 7 with LSM900).
Trf2 antibody
Manufactured by Abcam
The TRF2 antibody is a laboratory reagent used in scientific research. It is designed to detect and bind to the TRF2 protein, which is a component of the telomere complex. The TRF2 antibody can be utilized in various experimental techniques, such as Western blotting, immunoprecipitation, and immunofluorescence, to study the role and regulation of the TRF2 protein in cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Rabbit polyclonal anti γh2ax antibody, by Cell Signaling Technology (1 mentions)
Vectashield mounting medium, by Thermo Fisher Scientific (1 mentions)
Sin3a antibody, by Abcam (1 mentions)
Pierce agarose chip kit, by Thermo Fisher Scientific (1 mentions)
Power sybr green pcr master mix, by Thermo Fisher Scientific (1 mentions)
2 protocols using trf2 antibody
1
Immunofluorescence Analysis of Zona Pellucida-free Embryos
2
ChIP Assay for Telomere Protein Binding
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ChIP was performed using the Pierce™ Agarose ChIP Kit (Thermo Fisher Scientific, Inc.). Briefly, 1.5 × 107 cells were crosslinked in 1% formaldehyde, lysed and enzymatic digestion with Micrococcal nuclease (MNase). Chromatin was divided into equal amounts of immunoprecipitation with the TRF2 antibody (Abcam, ab13579), or rabbit IgG as negative control (Santa Cruz Biotechnology). Chromatin extracts were incubated on a rotator with 20 µl of ChIP Grade Protein A/G Plus Agarose for 3 h at 4 °C. Bound agarose beads were harvested by centrifugation (12.000 rpm, 15 s) and washed; the precipitated protein-DNA complexes were eluted from washed beads and incubated twice at 65 °C for 1.5 h with NaCl and Proteinase K to revert cross-links. Purified DNA was subjected to qPCR using Power SYBR™ Green PCR Master Mix (Thermo Fisher Scientific Inc.) and telomere primers (Table S2 ) as previously described [22 (link)]. Results are expressed as fold change as compared to IgG set as 1.
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