I am doing a gravity purification of 6x-His tagged staph nuclease in a pET28b+vector under native conditions, with no special conditions. This protein is very easy to purify and usually boatloads can be produced with the standard imidazole elution protocol. I want to order a fairly cost effective resin, as we will be using it for a summer academy with 8 groups of students. The protocol calls for 15 ml of resin (which I think is overkill). Any recommendations on the best and least expensive way to go about doing 8 column purifications with at least a 10 ml bed volume each? I am new to buying Ni-NTA resin and could use some suggestions on which types work best and which types of columns should be used. Thanks!
Although this post is now several years old it seems that that this topic is still relevant for many scientists. According to my experience the best Ni-NTA resins you can buy on the current market are from Cube Biotech (https://cube-biotech.com/products/purification-resins/his-affinity-resins/ni-nta-agarose/). They are less expensive compared to other resins and have the highest binding capacity on the market (80 mg/ml). I always get good results with them and can confidently recommend them to you.
I agree with Zdeno; if the protein is easy to purify, it is way more cost-effective to go with IDA (although you'll probably have to tweak the purification parameters).
At least in the UK, Pierce (ThermoScientific) sell a resin called HisPur Ni-NTA that as far as we can tell behaves identically to the Qiagen resin (in fact, it probably is manufactured by Qiagen and then rebranded by Thermo), but at a significant cost saving. For example, 100 ml HisPur Ni-NTA for £482.85 vs 100 ml Qiagen Ni-NTA for £667.44. I wouldn't go buying the more expensive superflow resins unless you need to pump at fairly high flow rates (e.g. superflow resins can go at easily 15 ml/min) - regular Ni-NTA is fine for gravity flow columns and low flow rates.
Also, as mentioned by Ermelinda, don't throw the used resin away, you can easily regnerate it and re-use many, many times. Example regeneration: Remove Ni2+ with 50 mM EDTA, wash with water, wash with 1M NaOH to remove remaining proteins, endotoxin and other stuff, then wash back with water until pH returns to neutral and store in 20% ethanol.
I'm pretty sure 15 ml is overkill too - 1 ml of resin typically binds 10-50 mg/ml protein. I think you could reduce costs substantially by reducing the amount of resin (and also reduce non-specific binding too).
look into the ligand density and then select a particular resin... depending upon the sample resin with high density/low density can be decided.......rule says if you want to purify protein in bulk and less specificity go for Ni-IDA and if you need more specificity than you need to choose metal ion other than Ni , you can try even Zn/Co on NTA
Qiagen and Roche brand were reliable and showed relatively good purity with 10mM imidazole co-treatment during Ni-NTA resin-sample incubation. Clontech brand looked horrible and showed lots of non-specific contaminants with the same condition. Perhaps that required their specific condition to purify His-tag proteins.
Although this post is now several years old it seems that that this topic is still relevant for many scientists. According to my experience the best Ni-NTA resins you can buy on the current market are from Cube Biotech (https://cube-biotech.com/products/purification-resins/his-affinity-resins/ni-nta-agarose/). They are less expensive compared to other resins and have the highest binding capacity on the market (80 mg/ml). I always get good results with them and can confidently recommend them to you.
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