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Gradient

It was demonstrated when dealing with peptide mapping that the solute s value, i.e. the degree by which a solute is affected by a change in the mobile phase polarity (gradient), is substantially affected by the solute molecular weight.

The use of broad range (5-100% organic/mobile phase B) gradients are never recommended, with shallower gradients over a much smaller polarity window being the norm; both chromatographic efficiency and selectivity of the Fc and Fab of Herceptin are affected by changing the gradient polarity range (Figure 1).

Effect of gradient polarity ranges on Herceptin Fc and Fab chromatography

Figure 1: Effect of gradient polarity ranges on Herceptin Fc and Fab chromatography

 
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