Blue dextran is used in affinity chromatography, gel filtration chromatography, protein chromatography and molecular weight markers. Blue dextran has been used to study Alzheimer′s disease .
What is the significance of using blue dextran in molecular exclusion chromatography when is it used and why?
Blue dextran is also used for checking the column packing . A symmetrical peak of elution indicates homogeneity of packing. The inner volume (Vi) of the column can be determined by subtracting the void volume from the elution volume of small molecules such as glucose or tyrosine having K d — 1.0.
How does gel filtration calculate void volume?
- Determine the void volume (Vo) by running a void marker and obtain the elution volume.
- Ascertain the total liquid volume (Vt) by running a total liquid volume marker and obtain the elution volume.
- Calculate the separating volume of the column (Vi) by subtracting the void volume from the total volume (Vt – Vo).
Is dextran a polymer?
Structure. Dextran is a polymer of anhydroglucose . It is composed of approximately 95% alpha-D-(1-6) linkages. ... Native dextran has been found to have a molecular weight (MW) in the range of 9 million to 500 million.
What is the meaning of exclusion limit?
In SEC, the exclusion limit is the molecular weight at the upper limit of a column's working range, beyond which molecules are too large to get trapped in the stationary phase and will elute together in the void volume of the column . Many SEC packings are referred to by their exclusion limit.
How do you calculate Kav?
Kav = ( Ve – Vo)/(Vc – Vo ) where Ve = elution volume, Vo = column void volume (7.71 mL based on Blue dextran elution volume), Vc = geometric column volume (24 mL).
How can I improve my gel filtration?
Increase in column length increases the resolution and increase in column diameter results in high bed volume and hence higher column capacity. The fractionation range and the exclusion limit can be controlled by varying pore size. The smaller the particle size of the gel, the higher the resolution achieved.
What is Sephadex g75?
General description. Sephadex ® G-75 is well-established gel filtration medium for desalting and buffer exchange of large biomolecules >80,000 molecular weight. Sephadex ® is a gel filtration medium prepared by crosslinking dextran with epichlorohydrin. ... Sephadex ® G-75 is for larger molecules.
How do you calculate gel filtration?
Gel Filtration. Ve = Elution volume (volume of solvent between injection and elution) . Dictated by proportion of porous matrix available to molecules (Kd).
Is dextran 70 safe for eyes?
This product (especially ointments) may temporarily cause blurred vision right after being placed in the eye(s). Do not drive , use machinery, or do any activity that requires clear vision until you are sure you can perform such activities safely. Consult your pharmacist or physician.
Is dextran A sugar?
Dextran is a branched polymer of dextrose (glucose) of very high molecular weight, belonging to the group of colloids.
Is dextran a protein?
18.2.
Dextran is a hydrophilic polysaccharide which is composed of α-1,6-linked d-glucopyranose (Fig. 18.7). ... Dextran is a biocompatible and nontoxic polysaccharide which is widely used in pharmaceutical and biomedical applications [23].
Why do large molecules elute first?
Smaller molecules experience a more complex pathway (like a maze) to exit the particle than do larger molecules. Because molecules that have a large size compared to the pore size of the stationary phase have very little entrance into the pores , these larger sized molecules elute first from the column.
What is the exclusion limit of the columns used in our experiment?
The smaller the molecules, the slower they move through the column. Molecules greater than 60,000 pass around the beads and are excluded from the column quickly – also referred to as the exclusion limit of a column.
What is gel filtration used for?
Gel filtration chromatography can be used to separate compounds such as small molecules, proteins, protein complexes, polysaccharides, and nucleic acids when in aqueous solution . When an organic solvent is used as the mobile phase, the process is instead referred to as gel permeation chromatography.
What is blue dextran?
Blue Dextran is a high-molecular-weight glucose polymer (original mol wt 2 x 10(6) g/mol) containing covalently bonded Reactive Blue 2 dye (approximately mmol/g dextran). This blue dye is known for its high binding affinity to a wide variety of proteins, with a particularly high affinity for serum albumin.
