UPLC-triple Quad for targeted quantitation

Acquity UPLC & Xevo TQ-S (Waters)

Plant Sciences group & RIKILT

twan.america@wur.nl

Xevo TQ-S

Acquity UPLC

Accurate and sensitive detection and quantitation of small biomolecules, such as:

• Metabolites, Hormones

• Toxins, elicitors

• Drugs, pesticides

• Peptides (proteins)

specific expertise for:

• development of targeted methods for protein quantitation based on proteomics marker analyses

• targeted detection of plant hormones

SRM / MRM principle

[Protein digest]

SPE sample prep

LC-MS separation

Precursor MS/MS

Fragment m/z

SRM transition

● Q1: precursor m/z

● Q2: fragmentation

● Q3: fragment m/z

Target extract SPE

Proteomics MS: peptide quantification

Compounds/Peptides elute from LC elution profile

MRM signal is integrated over peak area

Peak area is directly correlated to peptide load

Relative quantitation between samples

Synthetic peptides as (quantitative) reference

Absolute quantitation possible

Keywords

Targeted detection

● Selective

● Sensitive

● Quantitative (accurately)

● Broad concentration range

● Multiplexed

● Diverse

● Fast

Targets: Metabolites Hormones Toxins, elicitors Drugs, pesticides Peptides (proteins)

Method comparison

Untargeted LC-MSE

● Identification and (semi)-quantitation of a large number

of proteins in complex samples: thousands of peaks

● Optimise separation and resolution( long runs, ~2 /day)

● Highly detailed quantitative comparison

● Determine optimal marker peaks & fragments

● Transfer targeted method to QQQ

Targeted Triple Quadrupole MS (QQQ)

● Selected set of peptides

● Increased separation speed (typically 10min.)

● Increased sample throughput (up to 150/day)

LC-MS for identification and quantitation

Proteomics using LC-MS

Proteomics facility

PRI-Bioscience

twan.america@wur.nl