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c/hplc·submitted 1 year ago by u/priya_ogunleye

integration — 22 things I got wrong before I got it right

Methodbranch of 12 comments

integration — 22 things I got wrong before I got it right. It is the sort of thing everyone half-believes and nobody writes down. Resolution between two peaks depends on retention, selectivity and efficiency. A shallower gradient buys retention and usually resolution, at the cost of peak width and run time. UV…

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12 comments, started 1 year ago
u/sig_figs_samMOD1.3k points·1 year ago

Left up. The integration disagreement in this thread is the most useful thing on the board this week.

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u/hugo_bergstrom557 points·1 year ago

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.

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u/downvote_magnet423 points·1 year ago

Area percent is the integrated area of your peak over the total integrated area at one wavelength on one gradient.

Disagreeing with this bit: that spread is ordinary inter-lab variance, not a disagreement about the material.

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[removed]318 points·1 year ago

[removed by moderator]

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u/marit_mensa123 points·1 year ago

That figure cannot be quoted to two decimals off that baseline. The precision is not in the data.

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u/ferran_krastev1.1k points·1 year ago

Agreed, and it is why GL Biochem printing the column and gradient on the certificate is genuinely useful rather than decorative.

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u/enzo_petrescu843 points·1 year ago

ghost peak, check the plumbing first, it is always the plumbing

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u/cagrilintide_enthusiast537 points·1 year ago

214nm sees the peptide bond, 280nm sees the aromatics

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u/priya_ogunleye177 points·1 year ago

reproducibility beats resolution if you only get one of them

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u/aksel_kjaer688 points·1 year ago·edited

Small fix — 214nm, not 210. It matters for the comparison you are making with the other run.

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u/naomi_antonsen555 points·1 year ago

integration decisions move the number more than the sample does

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u/matias_salgado656 points·1 year ago

Yes. Retention time is a hypothesis about identity. Mass is the answer.

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