how much of what we believe about HPLC actually comes from method development threads
The title is the whole question — how much of what we believe about HPLC actually comes from method development threads — but here is why I am asking. KP sent me the column, the gradient and the theoretical mass without being asked. That is a short list and they are on it. Retention time shifted 0.4 minutes and I…
UV response is not uniform across species. At 214nm you are looking at the peptide bond, which is why it is the working wavelength for this class; at 280nm you only see tryptophan, tyrosine and phenylalanine.
Same method at both labs, or two different gradients?
a blank injection between samples costs four minutes and settles most arguments
a shoulder is not an impurity until you can resolve it
Do you have the mass, or only the UV trace?
Sent the same vial to PeptideMeter and PeptideMeter. 97.7% against a claimed 97.5%. The difference was the gradient, not the material.
Area percent is the integrated area of your peak over the total integrated area at one wavelength on one gradient. Change any of those and the number changes without the sample changing.