Users' questions

What is void volume in gel filtration chromatography?

What is void volume in gel filtration chromatography?

excluded volume
The void volume refers to the excluded volume i.e., the space between the particles. And the matrix volume refers to the solid component of the particles that fills the column bed. The pore diameter defines the exclusion limit of the gel.

What is void volume?

Void volume refers specifically to the volume of the liquid phase contained inside a column. The same term is sometimes also used informally to refer to the volume of a cavity in the column/tubing or fittings. Void volume is also known as dead volume.

How do you calculate void volume in size exclusion chromatography?

Void volume is often expressed as a percentage of the total column volume. For example, if the particles of stationary phase occupy 70% of the total column, the void volume would be 30% of the total column volume.

How do you calculate void volume in gel filtration?

  1. Determine the void volume (Vo) by running a void marker and obtain the elution volume.
  2. Ascertain the total liquid volume (Vt) by running a total liquid volume marker and obtain the elution volume.
  3. Calculate the separating volume of the column (Vi) by subtracting the void volume from the total volume (Vt – Vo).

What should the column size be for Sephadex G-25?

If the sample volume is very small a column of small size is recommended. If dilution of the sample must be kept to a minimum a small particle size gel filtration medium should be selected (e.g. Sephadex G-25 Fine or Sephadex G-25 Superfine). Pressure/Flow The particle size has a large influence on the pressure drop over the packed bed.

How to calculate the void volume of a column?

For columns with superficially porous packings, use the formula pi * (r) 2 * L * 0.49. This information is useful for chromatography methods involving a retentive mechanism like reverse phase or normal phase. The formulas shown above merely reflect an estimate of the void volume of a packed column (interstitial volume plus the pore volume).

What is the molecular mass exclusion limit for Sephadex G-25?

The pore size is chosen such that the large molecular weight solute is excluded from the gel matrix. Sephadex G-25 has a nominal molecular mass exclusion limit of 5000 for proteins [5].

What is the void volume in gel filtration?

Void volume is the volume of mobile phase (Vm or V0) in a column. In an ideal case, it is equal to the mobile phase hold-up volume. For example, if the stationary phase occupies 40% of the total column volume, the void volume would be 60% of the total column volume. Also, what is the principle of gel filtration?