Phenyl Focurose FF (HS)

Resin

Highly cross-linked 6% agarose

Particle size range 

45-165µm

Average particle size (D50)

90±5µm

pH stability

2-14 (long-term) 3-13 (short-term)

Chemical stability

Compatible with all common buffer solutions, 1M acetic acid, 1M sodium hydroxide, 8M urea, 6M guanidine hydrochloride, 30% isopropanol, and 70% ethanol

Linear flow velocity (0.3 MPa)

≥300 cm/h

Maximum pressure

≤0.3MPa

Storage solution

20% ethanol

Storage conditions

4℃-30℃

Influencing Factors

Mechanism

Recommendations

Ligand structure

Different ligands have varying binding affinities for proteins

It is advisable to perform pre-experiments to select the appropriate resin

Ligand concentration

Higher ligand concentrations result in stronger binding affinity

Pre-experiments should be conducted to determine the optimal ligand concentration

Sample properties

The hydrophobicity of a protein depends on the distribution of hydrophobic groups on its surface

/

Salt concentration

Higher salt concentrations lead to stronger binding between the ligand and the protein, but excessively high salt concentrations can cause protein precipitation

It is important to evaluate protein solubility and stability under different salt concentrations

Salt type

Different types of salts can yield different binding effects

Priority should be given to (NH₄)₂SO₄ and NaCl

Temperature

Higher temperatures enhance protein hydrophobicity

It is crucial to maintain a consistent temperature, preferably at room temperature

pH

Extreme pH values can affect protein solubility and stability, as well as binding efficiency

It is recommended to maintain the pH within the range of 5.0-8.5 while ensuring protein solubility and stability

Issue

Possible Causes

Solutions

Target protein does not bind or has low binding capacity during purification

Overloading of sample

Reduce the sample load

Sample flow rate is too fast

Lower the sample flow rate

Aggregation of impurities proteins or lipids in the resin

Clean the resin promptly and effectively or replace it with a new resin

Lower salt concentration in the equilibration buffer or weaker hydrophobicity of the target molecule

Increase the salt concentration in the equilibration buffer or change the type of salt, or replace it with a hydrophobic resin with stronger binding capacity

Not collecting the target protein during elution or collecting only a small amount of the target protein

Target protein does not bind to the resin or has low binding capacity

First, confirm if the target protein binds to the resin

Insufficient elution time

Reduce the flow rate to extend the retention time of the elution buffer

Insufficient elution volume

Increase the elution volume

Excessive binding strength between the target molecule and the resin

Reduce the salt concentration in the equilibration buffer or change the type of salt, or replace the hydrophobic resin with weaker binding capacity. Add additives (such as a small amount of detergent or low concentration organic reagents) to the elution buffer

Target protein purity is low

Sample not pre-processed

The sample must be centrifuged or filtered before loading onto the column

The sample has high viscosity

Dilute the sample with an appropriate equilibration buffer to reduce viscosity

Incomplete removal of impurities

Increase the washing volume until the baseline stabilizes and matches the equilibration buffer

Impurities such as proteins or lipids aggregate and precipitate in the resin

Clean the resin promptly and effectively

Poor elution conditions, such as excessive elution flow rate or steep gradient

Optimize the elution conditions

Poor column packing

Repack the column or purchase a new one

Large sample volume retained at the top of the column

Repack the column or reduce the sample volume

Inappropriate selectivity of the resin

Screen and select an appropriate hydrophobic resin

Microbial growth in the resin

After using the resin, store it correctly and promptly to prevent microbial growth.

Decrease in resin loading

Too fast sample flow rate

Reduce the sample flow rate

Aggregation of proteins or lipids in the resin resulting in decreased loading

Clean the resin promptly

Excessive use

Replace with a new resin

Rapid increase in chromatographic peak

Overly tight packing of the resin

Repack the column

Slow or tailing chromatographic peak

Loose packing of the resin

Repack the column

Cracks or dryness in the column bed

Leakage or introduction of large air bubbles

Check for leaks or bubbles in the tubing and repack the column if necessary

Slow liquid flow

Aggregation of proteins or lipids

Clean the resin or membrane promptly

Protein precipitation in the resin

Adjust the composition of the equilibration and elution buffers to maintain the stability of the target molecule

Microbial growth in the separation column

All reagents used must be filtered and degassed. The sample must be centrifuged or filtered before applying it to the column