Microscopy system with auto-focus adjustment by low-coherence interferometry

A method for automatic relative focus adjustment of an autofocus microscopy system that comprises a low coherence interferometer and a beam splitter that transmits light to a microscope and reflects light from the low coherence interferometer into an optical path parallel to an optical axis of an imaging objective, wherein a sample is covered by a cover material, the method comprising:

• computing A-scan data of a relative air/cover material position;
• computing A-scan data of a relative sample position;
• calibrating a focus position;
• computing a thickness of the cover material;
• processing a motor set position;
• and
• commanding a movement of a motor unit;
• wherein if a signal back scattered from the sample is less than 10 dB above a noise floor, storing a last position corresponding to a signal back scattered from the sample that is greater than 10 dB;
• wherein if a signal back scattered from the sample is less than 10 dB above the noise floor, the last position is used for the processing.

A method for operating a microscopy system that comprises a low coherence interferometer and a beam splitter that transmits light to a microscope and reflects light from the low coherence interferometer into an optical path parallel to an optical axis of an imaging objective, the method comprising:

• focusing light from the low coherence interferometer into a sample by the
• imaging objective, the sample being covered by a partially reflecting medium;
• providing to the low coherence interferometer a detection aperture that is not coincident with an aperture that is defined by an area with a point symmetry of an illumination aperture, the point symmetry being an intercept of the optical axis of the imaging objective with an aperture plane;
• and
• detecting a portion of back-scattered light from the sample and excluding a substantial portion of light reflected from the partially reflecting medium.