Christina mentions the histoneantibodies website, but after some experience with it, I no longer trust it as I cannot determine how they decide what the off targets are and what makes a good antibody. For example, they listed the Active Motif H3K27me3 antibody as being very poor with very strong off targets, but there's no evidence to back it up. When I looked at the databases they referenced, it passed all ChIP tests, and the western/dot blots looked good, especially in comparison to the millipore antibody, which the histone website listed as being better.
That said, I pick my antibodies by looking for ChIP-grade listed ones (abcam and active motif are good usually), and looking for their usage in publications, to see which are most trusted in the field. You should get monoclonal antibodies if possible, but most of them are mouse species, which makes them rather non-ideal to use if you are working with a mouse model as I am.
The first thing to proceed with is to know what is your target of interest in ChIP process. Once you select your target (e.g. TF, Histone methylation/acetylation etc.), there are plenty of high quality efficient Abs available commercially.
If you have problem in selecting Abs among multiple available commercially, then do not hesitate to contact seller's customer support.
Good Luck !
Hi,
Some Abs has been extensively used and validated by consortiums, e.g. Blue Print vhttp://www.blueprint-epigenome.eu/index.cfm?p=D6F8811F-DACF-7979-CEAC0B9034C28037
thank you himanshu and Irina for kind reply,
as i am working on mouse epigenetics, i am interested in Ab for histone ONLY, undoubtedly for most important and established histone marks playing role at developmental stages. i will go through with your link , but few points are still to be consider at best.
1. host in which Ab was raised
2. does the supplier will give standard primer set to have control during Chip ,like in case of Milipore?
3.because i am going to use fast chromatin protocol and specially at very low number of cells, so amount of Ab is also of my concern.
4. and the most importantly PRE-CLEARING STEP . should preclear dynabeads before use,or i need to clear Chromatin before incubating with Specific Ab?
thank u Leah Vandenbosch, i happy to get your reply with experience but please tell me how to look for off targates of an Ab before purchase. i want to go with Abcam but i dont think that they will give positive control primers and IgG along with the specific Ab like in case of Millipore.
thank you Christa Buecker-
thank you for your reply , i will be happy to know more about your experience with histone Ab ?
Hi,
In a case you are going to work with low amount of cell per ChIP, the following product can be interesting for you https://www.diagenode.com/files/application_notes/True_MicroChIP_and_MicroPlex_kits_Application_Note.pdf.
This kit has been developed and validated on 10.000 cells with many histones marks using Diagenode Abs. The shown data are made on human cells but a most part of Abs works on mouse cells and some mouse control primers are available. You can contact [email protected] if you are interested.
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