Quantitative proteomics (TMT and iTRAQ)
How TMT and iTRAQ measure thousands of proteins at once, applied to heart-valve development.
Role Researcher and presenterWhen Graduate coursework, UW-MadisonTeam Two authors
This is the oldest project on the page and comes from my science background. I include it because it shows how I learn a complicated technical field well enough to explain it to others.
The problem
Proteins do most of the work in a cell, and diseases often start with a protein being made too much or too little. Quantitative proteomics measures how much of each protein a sample contains, and TMT and iTRAQ are the standard ways to compare many samples in one experiment. Both attach chemical tags that look the same to the instrument until the sample is broken apart in the mass spectrometer, so a single run can measure several conditions side by side.
What I covered
My co-author and I built a 74-slide talk that took the audience through the whole workflow: extracting proteins, cutting them into peptides with trypsin, labeling each sample with a different tag, mixing the samples, running liquid chromatography and mass spectrometry, and using software such as Proteome Discoverer to identify and quantify proteins from the mass-to-charge data. We compared iTRAQ with TMT and spelled out where each method breaks down.
Applied to the heart
For the applied part, I looked at congenital heart defects. The study I presented used TMT mass spectrometry in zebrafish to see how the microRNA miR-21 shapes the formation of heart valves. The team identified over 250 proteins affected by miR-21, and bioinformatics and luciferase assays confirmed some of them as direct regulatory targets, which pointed to new leads on how valves form.
What carries over
Proteomics taught me to ask how a number was measured, and what could distort it, before I trust it. I do the same when I read an AI evaluation.
The presentation
