ChIP Anti-bodies CTCF

In ChIP, the most vital step is the binding of an antibody and choosing the right antibody. The binding affinity of different types of immunoglobulins to protein A or G differs significantly. Henceforth, it is recommended to choose either protein A or protein G coated beads. If you do not see any product in the positive control, add 5–10 μg of chromatin and 1–5 μg of antibody to each IP reaction and incubate with antibody overnight and an additional 2 hr after adding Protein G/A beads. If no product is observed in the experimental sample, add more DNA to the PCR reaction or increase the number of amplification cycles. Furthermore, if you have any problem with antibodies, make sure to use the ChIP-validated antibody.

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Found 3 matching solutions for this experiment

CTCF (D31H2) XP® Rabbit mAb #3418

Cell Signaling Technology

Upstream tips
-It is highly critical that the chromatin is of appropriate size and concentration.
Protocol tips
-For optimal ChIP and ChIP-seq results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP.
-It is important to keep the tissue cold to avoid protein degradation.
-Use fresh formaldehyde that is not past the manufacturer's expiration date.
Downstream tips
-Once in solution, store 1M DTT at -20°C.
Anti-CTCF Antibody

Merck Millipore

Protocol tips
ChIP-exo was performed as previously described6,20 with chromatin extracted from 50 million cells, ProteinG MagSepharose resin (GE Healthcare), and 5 ug of antibody. Two biological replicates were prepared for each ChIP target. To enable future comparisons to drug treatment experiments, cells were treated with vehicle (final 0.03% DMSO (dimethyl sulfoxide)) for 4 h prior to harvest.
Upstream tips
-Add protease inhibitors to all lysis solutions before use.
Protocol tips
-Use at an assay dependent concentration.
-Keep cells on ice between the rounds of homogenisations.
- Increase or decrease the homogenization step to maximize the yield of nuclei depending on cell line.
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