The largest database of trusted experimental protocols

Hitrap blue hp column

Manufactured by GE Healthcare
Sourced in Japan

The HiTrap Blue HP column is a pre-packed affinity chromatography column designed for the purification of a variety of proteins, including enzymes, hormones, and other biomolecules. The column contains a ligand that selectively binds to target proteins, allowing for their separation and purification from complex mixtures. The column is suitable for use in both small-scale and large-scale applications.

Automatically generated - may contain errors

5 protocols using hitrap blue hp column

1

Affinity Chromatography of ASP0-50%

Check if the same lab product or an alternative is used in the 5 most similar protocols
ASP0-50% was dissolved in approximately 1 ml of 20 mM HEPES buffer (pH 7.4), and dialyzed against 20 mM HEPES buffer (pH 7.4). After dialysis, sample (total protein amount of 42 mg) was applied to a HiTrap Blue HP column (5 ml, GE Healthcare, Tokyo, Japan) equilibrated with 20 mM HEPES buffer (pH7.4) using a manual syringe. After a 50-ml wash with 20 mM HEPES buffer (pH 7.4), the elution of proteins bound to the column was performed with a step-wise elution with 20 mM HEPES buffer (pH 7.4) containing 2 M NaCl, and 1 ml of fractions were collected. Aliquot of fractions from the column unbound (Blue pass) and bound elutes (Blue elute) were subjected to the AOND assay and protein content analysis (S1 Fig).
+ Open protocol
+ Expand
2

Wnt3a Purification from Conditioned Media

Check if the same lab product or an alternative is used in the 5 most similar protocols
Processed CM was applied to a 5 ml bed volume HiTrap Blue HP column (GE Healthcare). The column was protected from residual precipitate by an inline 0.45 μm syringe filter. The sample was applied to the column at ~2 ml/min with a peristaltic pump. The column eluate was collected during loading and α-Wnt3a immunoblot analysis performed to monitor Wnt3a binding to the column matrix. After loading up to 5 L processed CM and washing with 40 ml 20 mM Tris HCl, pH 7.45 containing 0.14 M NaCl and 1 % (w/v) CHAPS detergent (buffer A) the column was placed in an AKTA Prime FPLC system (GE Healthcare). All operations to this point were carried out at 4 °C. The column was washed with 150 ml buffer A at 2.5 ml/min and protein eluted with 1.5 M sodium chloride, 20 mM Tris-HCl, 1% CHAPS, pH 7.45 (buffer B) using the following gradient: 20 ml to 25% B; 5 ml to 35% B; 2.5 ml to 100% B followed by a further 75 ml at 100% B (flow rate = 2.5 ml/min). The collected fractions were pooled based on A280 and SDS-PAGE analysis.
+ Open protocol
+ Expand
3

Purification of Recombinant Erythropoietin

Check if the same lab product or an alternative is used in the 5 most similar protocols
The purified pcDNA 3.3 expression vector containing a sequence encoding EPO-WT or EPO mutant was transiently transfected into FreeStyle 293-F cell using 25 kD linear polyethylenimine (Polysciences). After 6 days, EPO protein was purified from culture supernatant using Hitrap Blue HP column (GE Healthcare), eluted with 1.5 M NaCl, and buffer exchanged into 20 mM Tris-HCI pH 8.45. Next, sample was loaded onto Hitrap Q HP column (GE Healthcare). The column was washed with 20 mM sodium acetate pH 4.00 followed by second wash with 20 mM Tris. EPO protein was then eluted with 1 M NaCl. Purified EPO protein was buffer exchanged into a storage buffer (50 mM sodium phosphate buffer, pH 7.0, 1.5% glycine and 0.003% tween-20).
+ Open protocol
+ Expand
4

Cortisone and 7α-Hydroxycholest-4-en-3-one Synthesis

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cortisone and 7α-hydroxycholest-4-en-3-one were purchased from Steraloids. Finasteride was purchased from Sigma-Aldrich. NADPH was purchased from EMD. NADP+ was purchased from Roche Applied Science. E. coli strain C41 (DE3) was provided by Dr. J. E. Walker (Medical Research Council Laboratory of Molecular Biology, Cambridge, UK). HisTrap Fast Flow column (5 mL) and HiTrap Blue HP column (5 mL) were purchased from GE Healthcare. All other reagents were of American Chemical Society quality or higher.
+ Open protocol
+ Expand
5

Protein Purification and Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols
General chemicals and the phosphate assay kit (MAK308) were from Sigma-Aldrich. Restriction enzymes and commercial apyrase were from New England Biolabs. Chelating Sepharose fast flow resin and the pre-packed HiTrapBlue HP column were from GE Healthcare or Cytiva. Ni-NTA agarose resin was from Qiagen. Coelenterazine H BRET substrate was from Nanolight. Dialysis membrane was from Spectrum Labs.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!