The words of this tutorial, step by step. The text lives in src/smappy/tutorial/topics/; the directions it follows are in src/smappy/tutorial/STYLE.md.
1. card: Drift correction: RCC and COMET
Over the minutes of an acquisition the sample moves, by tens to hundreds of nanometres. Each localization is off by where the sample was in its frame, and the picture blurs.
Drift is estimated from the localizations themselves, then subtracted: no beads needed.
spoken: Over an acquisition the sample drifts, and the picture blurs. SMAPpy estimates the drift from the data and takes it out.
2. To practise, Simulate adds drift on request, and remembers how much, so the correction can be checked.
3. The first half of the acquisition in red, the second in green. Without drift they would overlap in yellow; here they do not.
4. card: RCC and COMET
RCC renders the data in a few time windows and cross-correlates them. It is fast and robust, and limited by its rendering.
COMET fits the drift to the localizations themselves, finer and slower. Before a long run it says how long, and offers RCC first.
spoken: Two methods: RCC correlates rendered time windows, fast and robust. COMET fits the localizations themselves, finer.
5. A drift plugin estimates from what the current layer shows, so give it all the frames back first: one layer, no frame filter.
6. RCC's main settings: how many time windows, the pixel of the rendering, and how far the sample may have moved.
7. The drift it found, in x, y and z over the acquisition. It is subtracted from every localization.
8. Red and green now overlap in yellow: the two halves agree again.
9. To try COMET on the same data, undo the correction first: File, Undo, or Ctrl+Z.
10. COMET's settings: the window size, how far the sample may have moved, and a spline that smooths the drift between windows.
11. On a large dataset COMET says how long it will take before it starts, and offers RCC first: seconds for the bulk of the drift.
12. COMET's estimate of the same drift. On real data it is often the finer of the two; RCC first is a good way to start a long run.
13. The drift curve is saved with the file, so Plot draws it again after the file is reopened, and the correction can be undone.
14. card: What to remember
- See drift with two layers, early and late frames, in two colours.
- RCC: fast and robust. COMET: finer, and it warns before a long run.
- Correct drift before measuring; undo brings the old table back.
spoken: That is drift correction: see it in two colours, correct it with RCC or COMET, and measure after.