Three-dimensional resolution-enhancement divided aperture correlation-differential confocal microscopy with nanometer axial focusing capability

Abstract

Divided aperture confocal microscopy (DACM) provides an improved imaging depth, imaging contrast, and working distance at the expense of spatial resolution. Here, we present a new method-divided aperture correlation-differential confocal microscopy (DACDCM) to improve the DACM resolution and the focusing capability, without changing the DACM configuration. DACDCM divides the DACM image spot into two round regions symmetrical about the optical axis. Then the light intensity signals received simultaneously from two round regions by a charge-coupled device (CCD) are processed by correlation manipulation and differential subtraction to improve the DACM spatial resolution and axial focusing capability, respectively. Theoretical analysis and preliminary experiments indicate that, for the excitation wavelength of λ = 632.8 nm, numerical aperture NA = 0.8, and normalized offset v = 3.2 of the two regions, the DACDCM resolution is improved by 32.5% and 43.1% in the x and z directions, simultaneously, compared with that of the DACM. The axial focusing resolution used for the sample surface profile imaging was also significantly improved to 2 nm.

Annular-beam dual-detection confocal reflectance microscopy for high-speed three-dimensional surface profiling with an extended volume

“…To overcome this limitation, researchers have tried to eliminate the repetitive two-dimensional scanning process through approaches such as chromatic confocal microscopy (CCM), differential confocal microscopy (DCM), and divided-aperture differ ential confocal microscopy (DDCM). CCM, DCM, and DDCM encode the height information so that height can be calculated from the signal without axial scanning [9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24]. First, CCM encodes the height of the sample surface spectrally in the axial direction.…”

“…DDCM combines the divided-aperture confocal microscopy and DCM. In other words, it uses the pupil plane of the objective lens, which is divided into illumination and collection to move the airy disc transversely on the detector plane when the object is translated axially, and it measures intensities with two point detectors that have a transverse offset from the optical axis because the linear relationship between the height of the sample surface and the differential signal can be used as a lookup table [21][22][23][24].…”