Title: Dual-LED-based multichannel microscopy for whole-slide multiplane, multispectral, and phase imaging,
Abstract: We report the development of a multichannel microscopy for whole-slide
multiplane, multispectral, and phase imaging. We use trinocular heads to split
the beam path into 6 independent channels and employ a camera array for
parallel data acquisition, achieving a maximum data throughput of ~1 gigapixel
per second. To perform single-frame rapid autofocusing, we place two
near-infrared LEDs at the back focal plane of the condenser lens to illuminate
the sample from two different incident angles. A hot mirror is used to direct
the near-infrared light to an autofocusing camera. For multiplane whole slide
imaging (WSI), we acquire 6 different focal planes of a thick specimen
simultaneously. For multispectral WSI, we relay the 6 independent image planes
to the same focal position and simultaneously acquire information at 6 spectral
bands. For whole-slide phase imaging, we acquire images at 3 focal positions
simultaneously and use the transport-of-intensity equation to recover the phase
information. We also provide an open-source design to further increase the
number of channels from 6 to 15. The reported platform provides a simple
solution for multiplexed fluorescence imaging and multimodal WSI. Acquiring an
instant focal stack without z-scanning may also enable fast 3D dynamic tracking
of various biological samples.