Quadrupole, time-of-flight, orbital trap: what each class can and cannot see
Every instrument specification quoted in an advertisement is a best case obtained on a calibration mixture, not on a submitted vial.
HPLC
Page 4 of 5 of this archive, newest first.
Peak capacity, and the honest measure of a separation
Column chemistry, particle size and pore diameter determine what the separation is capable of before the gradient is even programmed.
Before anything is weighed, the peptide has to be charged
A brief and unromantic tour of the interface between a liquid sample and a vacuum.
Two ways to lift a peptide into a vacuum, and why the answer differs
The ionisation method determines the charge states you see, the adducts you must account for, and the modifications you might destroy in the process.
Two gradients, one sample, two purities
A shallow gradient resolves impurities that a steep one runs into the parent peak. Both methods are legitimate; only one of them can see the small stuff.
Electrospray, MALDI, and the choice that shapes every spectrum after it
A brief and unromantic tour of the interface between a liquid sample and a vacuum.
Pore size, particle size, and why peptides need the wide pores
A shallow gradient resolves impurities that a steep one runs into the parent peak. Both methods are legitimate; only one of them can see the small stuff.
What competing on one number does to a market
A market that competes on one measurement will optimise that measurement. There are legitimate ways to raise a purity figure that involve no change to the product at all.
What “LC-MS: conforms” tells a reader, which is almost nothing
What the Journal asks for when it writes to a supplier about an identity claim, and how often it gets it.
The figure of merit this trade chose, and the four it did not
Chromatographic purity is cheap, fast and comparable-looking. Those three properties, and not its usefulness, explain why it became the industry’s single figure of merit.
Quadrupole, time-of-flight, orbital trap: what each class can and cannot see
Every instrument specification quoted in an advertisement is a best case obtained on a calibration mixture, not on a submitted vial.
Two analysts, one chromatogram, two answers
We work through a single chromatogram twice, under two integration conventions, and show where the difference comes from.
Where the solvent front ends is a decision, not a fact
We work through a single chromatogram twice, under two integration conventions, and show where the difference comes from.
The isotope pattern tells you the charge, and the charge tells you the mass
A peptide has a monoisotopic mass and an average mass, they differ by several daltons at this molecular size, and a certificate that does not say which it quotes cannot be…
Manual reintegration is legitimate, and it should be declared
We work through a single chromatogram twice, under two integration conventions, and show where the difference comes from.
The number at the bottom of the certificate
A purity percentage is the most quoted figure in the research-peptide trade and one of the least examined. We asked four laboratories to analyse the same vial and got four…
Equal area is not equal mass: the response factor problem
Peptide bonds absorb strongly near 214 nm; aromatic side chains absorb near 280 nm. A method reading at 280 is blind to any fragment lacking an aromatic residue.
The impurity hiding underneath the parent peak
The most useful single addition to any purity determination is a second separation under a different pH or on a different stationary phase, and the second-most useful is a…
D-amino acids are invisible to mass spectrometry, and that is the whole problem
What would be needed to catch each of these, and what it would cost.
The specification sheet says 500,000. The report says conforms.
Calibration drift is real, unremarkable, and the reason serious laboratories run internal standards.