GATTAquant microscopy standards

February 13, 2015 at 5:27 pm | | review, single molecules

Jürgen Schmied from GATTAquant came by the other day and let me play around with some of their cool DNA origami fluorescence standards.

The PAINT sample was really cool. It has short oligos on the DNA origami and complementary strands labeled with dyes in solution. The binding/bleaching kinetics are such that each hotspot blinks many times during an acquisition. After a quick 10,000 frame acquisition over 3 min, we collected a dataset that we could easily get a super-resolution image. We used ThunderSTORM to fit the data and correct for drift. But without any other corrections, we could easily resolve the three PAINT hotspots on each DNA origami:

Screen Shot 2015-02-13 at 5.12.44 PM

But my favorite sample was actually the confocal test slide. It had two sets of dyes about 350 nm apart permanently labeled on each DNA origami.

Screen Shot 2015-02-13 at 5.13.05 PM

This let me test the resolution and image quality using different configurations on our Diskovery confocal/TIRF system.

Screen Shot 2015-02-13 at 5.13.21 PM

Each spot contained only about 4-8 dyes. So it was a much greater challenge to our microscope than TetraSpeck beads.

Screen Shot 2015-02-13 at 5.12.55 PM

I highly recommend GATTAquant test samples. Very fun.

UPDATE: Jürgen ran my data though GATTAquant’s analysis software and sent me the results below.


export3_FWHM histogram

export3_Distance histogram

a new PALM software package

February 13, 2015 at 2:11 pm | | nerd, software

2015-02-13 13.56.48

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