ChIP Anti-bodies GATA3

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 2 matching solutions for this experiment

GATA-3 Antibody (HG3-31): sc-268

Santa Cruz Biotechnology

Protocol tips
-Investigators may wish to consider using the primary antibody conjugated to sepharose or magnetic beads as an alternative to using secondary immunoprecipitation reagents (e.g., Protein A-Agarose) as described here.
Combining primary antibodies directed to different epitopes of the same protein may be advantageous in some cases.
Upstream tips
-Add protease inhibitors to all lysis solutions before use.
Protocol tips
-Use 2 µg for 25 µg of chromatin.
-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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