rapid-af

Real-time image-based autofocus for all wide-field optical microscopy systems

RAPID (Rapid Autofocusing via Pupil-split Image phase Detection) is an implementation of the phase detection principle for defocus measurement, applicable to all wide-field optical microscopes including light-sheet ones. In RAPID, two images of the same field of view are formed selecting the rays passing through two distinct portions of the system pupil. The mutual displacement between these images (the 'phase') is linearly proportional to the defocus.

The details of the optical setup can be found in

Silvestri, L., et al. "Universal autofocus for quantitative volumetric microscopy of whole mouse brains." Nature Methods 18.8 (2021): 953-958. https://doi.org/10.25493/AV5J-M46.

The code reported here, exploiting OpenCV, can be used to implement efficient image registration between the two pupil-split images.

What this code does

It performs optional image pre-processing, and computes cross-correlation to find mutual image displacement. This is the most general part of the RAPID concept, which is independent from the actual optical implementation.

What this code does NOT

It does not include packages for image grabbing (or camera management in general). Also, no code for actual defocus correction (e.g. by moving the detection objective) is provided. These modules depend on the hardware used in the microscope.

How to build this project

On Ubuntu, install the following packages:

sudo apt install cmake build-essential libopencv-dev

Create a build directory outside the current repository, then run cmake:

mkdir build
cd build

cmake /path/to/rapid/repository
make
sudo make install

This will install a binary named rapid-af and a library named librapid-af.so.

How to use rapid-af

In your CMakeLists.txt, add the following:

find_package(rapid-af REQUIRED)

Then, when linking libraries to your executable:

add_executable(myexecutable main.cpp)
target_link_libraries(myexecutable
    rapid-af
)

In your source file:

#include <rapid-af.h>

void main() {
    cv::Mat i1, i2;
    bool ok;
    const struct rapid_af::Options opt;
    cv::Point2f shift = rapid_af::align(i1, i2, opt, &ok);
}

Documentation

https://lens-biophotonics.github.io/RAPID-AF