Question

Explain how an antibody could be useful for purifying a protein and for determining its concentration.

Short answer

The proteins bind to antibodies at sites other than the antigen binding domain, an immobilized form of the proteins is used in purification. It involves selective enrichment or specific isolation of antibodies from serum. Protein concentration can be estimated by measuring the UV absorbance at 280nm; a strong peak shows here due to Tryptophan or Tyrosine residues. This can be readily converted into protein concentration using the Beer-Lambert law.

Step-by-step solution

Physiochemical fractionation

A separation process in which a certain quantity of a mixture like gas, solid, liquid, or enzymes is divided during a phase transition into several tiny fractions in which the composition differs according to a gradient.

Class-specific affinity

This type of affinity allows for purifying a selected class of immunoglobulin and discarding the unwanted types. A particular antibody called IgG binds to solid-phase by immobilized biological ligands (proteins, lectins, etc.)

Antigen-specific affinity

This is based on affinity purification in which antibodies in a sample bind to a particular antigen molecule through their specific antigen-binding domain. It purifies all antibodies that bind to the antigen without considering antibody class or isotypes.

Beer-Lambert law

It states that there is a linear relationship between the concentration and the absorbance of the suspension.

A = ɛcl

Where,

A = absorbance

ɛ = molar absorptivity

c = concentration

l = length of the light path

Hence, antibody can be used to purify proteins.