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Questions related to Protein Purification Techniques
I have a problem with my research about xylanase purification. When I precipitate the production media (protein sample) with crystal ammonium sulfate 60% saturation and I got the precipitat. After...
08 March 2020 8,940 6 View
Hello everyone, I'm trying to purify his-tagged protein under denaturing conditions using Guanidine Hydrochloride (Protocol from GE Healthcare-attached below). Though I'm getting my protein in...
12 December 2019 716 3 View
11 December 2019 8,833 3 View
I am conjugating Amyloid-beta (Abeta) peptide (MW ~4500) with a small molecule (starting with monomeric abeta). I tried purifying the product with protein concentrator with 3000 molecular weight...
15 August 2019 6,746 5 View
08 August 2019 3,245 6 View
Hello, I am trying to dialyze a protein I'm working with in a buffer containing 100 mM L-cysteine. From what I've read, L-cysteine should be dissolved in a low pH solvent, in an airtight...
02 July 2019 7,730 4 View
I routinely purify FLAG-tagged protein-RNA complexes that have been crosslinked with formaldehyde using FLAG peptide. Alternatively, it is possible to elute proteins from FLAG resins using high...
04 March 2019 6,770 3 View
I have protein tagged with mEOS3 and want to check it's concentration after purification with spectrophotometer. How can I do that? What wavelength I should use and what Extinction coefficient to...
01 January 2019 8,952 2 View
Am trying to purify my protein in Tris-Hcl pH 8.0 with DTT. I am keeping the protein for overnight dialysis. I found that, around this pH 8.5 in phosphate buffer stability of DTT is about 1.4...
31 December 2018 6,493 11 View
Hello, I am doing a research project on the production of purified protein derivative used in TB tests. From my research there are many methods but not enough information on what one particular...
11 November 2018 4,720 0 View
Can we store pierce glutathione agarose beads in 50 mM tris, 150 mM NaCl,pH 8.o after regeneration?
01 November 2018 4,547 3 View
I took a sample from a Mtb culture that is growing in Middlebrook 7h9 enriched with OADC(oleic acid,albumin, dextrose and catalase). The bacteria were harvested by centrifugation, and the...
15 March 2018 5,036 2 View
My protein (64kDa) expressed in insoluble inclusion bodies. I tried to purify it by ion-exchange and affinity but unable to purify protein. So, want to try ammonium sulfate precipitation of...
07 March 2018 749 4 View
03 March 2018 4,654 2 View
I did a trial run of a his-purification and found that my protein would be a little dilute for my purposes. Any thoughts on the best way to concentrate a denatured protein? This is my buffer and...
17 October 2017 5,488 7 View
Using PEG/salt aqueous two phase extraction system, after the partitioning of protein to either the polymer or salt phase, which are the suitable methods for concentrating of these proteins? Any...
09 October 2017 9,122 14 View
I have cultured primary enterocytes with lipids. After then I collected the medium and cells' microsome lumenal proteins for NaBr gradient (d=1.006~1.10), and each fraction (1ml) was collected...
31 July 2017 7,904 5 View
Hello everyone, I am trying to purify my protein (enzyme) expressed in Pichia pastoris through ion exchange chromatography. The protein is extracellular, so I extract the supernatant after...
28 July 2017 4,191 5 View
Hello, I have a problem during purifying my two proteins using Ni beads (GE healthcare). These two small proteins (about 6 kDa) were expressed in E. coli with an N-terminal His-SUMO tag. When I...
24 July 2017 1,788 3 View
Hello I have been trying to express and purify two truncated his-tag proteins(45KDa and 65KDa size without his-tag) using baculovirus expression system. I was however able to express the proteins...
22 July 2017 7,614 7 View
Hello.I have purified my protein with Ni-NTA coulmn and i have run my purified fractions on SDS-PAGE but i don't know how to calculate the purity of may bands. how should i do this?Thanks in advance.
18 July 2017 8,135 6 View
Hi! My target protein does not bind to the column. Before purification, I have confirmed that it has a His-tag by Western blot. Also, the protein is in the insoluble fraction so I carried out...
14 July 2017 5,552 10 View
The salts should be removed from the solution contains proteins and salts (calcium oxide/ sodium chloride) by diafiltration process. My question is 1) Can we use the ultrafiltration membrane for...
07 July 2017 7,989 9 View
Dear sir/madam I have been trying to purify protease enzyme using HIC column chromatography. But, the protein concentration was very less compare than crude protein. I loaded the...
07 July 2017 2,261 9 View