A perfect mix: DirectOverlay™ combines AFM and optical microscopy precisely and easily

The optical calibration and import feature DirectOverlay™ set a milestone in AFM. Invented by JPK Instruments in 2006, it revolutionized the way AFM and optical microscopy could be combined. Using AFM simultaneously with optical microscopy provides complementary information from the sample. Interesting locations on the sample can be identified with a wide variety of contrast methods such as optical phase contrast, DIC or VAREL contrast.

Additionally fluorescence techniques such as epi-fluorescence, confocal microscopy, TIRF, FRET, FCS etc. give insights about the behaviour or location of particular molecules. Combining AFM imaging or force measurements with these optical methods on the same spot at the same time is not straight forward.

First of all the optical image acquisition must be independent from the AFM image recording. A tip scanning AFM design is crucial for this. But a simple overlay of AFM image data and optical images afterwards shows a mismatch of both data sets, caused by errors in the optical lens systems.

These are not made for metrology measurements. To overcome this JPK developed the patented DirectOverlay™ software feature.

DirectOverlay™ examples

Click on the images for details

With this the user has:

• Perfect overlay of optical and AFM data with sub-diffraction limit precision with a click
• Direct "in optical image" selection of AFM measurements (imaging and force curves)
• Dramatic reduction of overview image scanning in AFM, giving faster results and lower tip contamination
• Optical image navigation to specific regions of interest, even without AFM scanning. This protects functionalized tips for molecular recognition, avoiding tip passivation from image scanning before the force measurements.