Measuring Small-molecule Inhibition of Protein Interactions in Live Cells Using FLIM-FRET
Bibliographic record
Abstract
This protocol was designed to quantitatively measure small-molecule displacement of proteins in live mammalian cells using fluorescence lifetime imaging microscopy-Förster resonance energy transfer (FLIM-FRET). Tumour cell survival is often dependent on anti-apoptotic proteins, which bind to and inhibit pro-apoptotic proteins, thus preventing apoptosis. Small-molecule inhibitors that selectively target these proteins (termed BH3-mimetics) are therefore a promising avenue for the treatment of several cancers. Previous techniques used to study the efficacy of these drugs often use truncated versions of both pro- and anti-apoptotic proteins, as they are membrane bound and hydrophobic in nature. As a result, the true efficacy of these drugs to displace full-length pro-apoptotic proteins in their native environment within a cell is poorly understood. This protocol describes FLIM-FRET methods to directly measure the displacement (or lack of displacement) of full-length Bcl-2 family proteins in live mammalian cells.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".