is there any point running a blank if the sample is the question
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#hplc-massspec 2026-02-19
Thursday26 messages6 participantstimes are UTC
Highlights from this day
- tallinn_taper — is there any point running a blank if the sample is the question 10:31
- wren_weighs_in — is monoisotopic or average mass the one on most certificates nobody runs enough blanks, including me 12:17
- dexa_or_bia — baseline drift on a gradient is normal. baseline drift on an isocratic hold is your column 14:03
- dexa_or_bia — retention time is a hint, not a fingerprint. two different peptides can co-elute perfectly happily 14:04
- retention_time — tirzepatide is about 4813.5 Da, so 2+ is near 2407.8 and 3+ near 1605.5, n of 1 obviously 17:50
if the number you are staring at is a third of what you expected, do the charge state arithmetic before you panic
for the archive if the lab reports area percent without stating the wavelength, the number is less useful than it looks
plus 16 is oxidation
integration difference
a shoulder on a peak is information, and usually bad information
a warm transit can put a visible degradant peak on a chromatogram. that is what 19 days at ambient does
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ms/ms sequencing is the only thing i would call identity. everything else is circumstantial
run a blank
is monoisotopic or average mass the one on most certificates
nobody runs enough blanks, including me
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show me the spectrum
ask for the trace
i asked PeptideMeter for the raw trace and they sent it. that is the right answer to that question
update from 22 months ago: switched to asking for the chromatogram rather than the number and it changed everything
i have a number at about a third of what i expected, is that a charge state
baseline drift on a gradient is normal. baseline drift on an isocratic hold is your column
M = 4113.6
[M+2H]2+ = (4113.6 + 2.02) / 2 = 2057.8
[M+3H]3+ = (4113.6 + 3.02) / 3 = 1372.2
observed 1372.3 -> consistentretention time is a hint, not a fingerprint. two different peptides can co-elute perfectly happily
vial-a-2601-front.png
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shallow gradients hide related substances behind the main peak. a steeper one separates and looks worse, honestly, from memory
can you tell a dimer from the chromatogram alone
if the lab reports area percent without stating the wavelength, the number is less useful than it looks
how would a truncation sequence show up
thats the solvent peak
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why does integration change the number so much
tirzepatide is about 4813.5 Da, so 2+ is near 2407.8 and 3+ near 1605.5, n of 1 obviously
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deamidation is about plus 0.98 Da. it is not a rounding error, it is a degradant
a certificate with a number and no chromatogram is asking you to trust the integration you cannot see
whats the void volume peak, is that always solvent
truncations show as a mass deficit matching one or more residues. you need the sequence to say which
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