Digital holographic microscope

Microscope comprising a light source producing a collimated light beam, a light beam guide system comprising a beam splitter configured to split the light beam into a reference beam and a sample illumination beam passing through the light beam guide system directed by at least one direction change mirror being pivotally actionable (TM1, TM2, TM3, TM4) to guide the reference beam and sample beam along their respective optical paths, a sample observation zone configured to receive a sample to be observed in a path of the sample illumination beam, a sample illumination device configured to direct the sample illumination beam through the sample observation zone and into a microscope objective, a beam reuniter configured to reunite the reference beam and sample illumination beam after passage of the sample illumination beam through the sample observation zone, a light sensing system configured to retrieve at least phase and intensity values of the light beam downstream of the beam reuniter, an optical path difference (OPD) adjustment device positioned in an optical path of the reference beam, the OPD adjustment device configured to adjust the optical path length of the reference beam relative to the sample illumination beam, and a control system, wherein the OPD adjustment device comprises a first pivotally adjustable light deviating element and a second pivotally adjustable light deviating element, each comprising a transparent or reflective material positioned in the optical path of the reference beam, whereby angles of inclination of the light deviating elements are controlled by the control system to adjust the optical path difference.

Method of controlling a microscope comprising a light source producing a light beam, a light beam guide system comprising a beam splitter configured to split the light beam into a reference beam and a sample illumination beam passing through the light beam guide system directed by at least one direction change mirror being pivotally actionable (TM1, TM2, TM3, TM4) to guide the reference beam and sample beam along their respective optical paths, a sample observation zone configured to receive a sample to be observed in a path of the sample illumination beam, a beam reuniter configured to reunite the reference beam and sample illumination beam after passage of the sample illumination beam through the sample observation zone, a light sensing system configured to retrieve at least phase and intensity values of the light beam downstream of the beam reuniter, an electronic control system, and a sample illumination device configured to direct the sample illumination beam through the sample observation zone and into a microscope objective, the sample illumination device comprising a mirror system configured to direct the sample illumination beam at a non zero illumination angle (α) with respect to an optical axis (A) of the microscope objective and a rotating beam system configured to rotate the angled sample illumination beam around the optical axis, the method wherein generating a look-up table (LUT) in a memory of the control system for adjusting an optical path length (OPL) of the reference beam by the control system of the microscope to keep an optical path difference (OPD) between the reference and sample beams below a coherence length of the light source, comprising the following steps:

• a) position the rotating beam system in a first position,
• b) position said at least one pivotally actionable direction change mirror (TM3, TM4) configured to direct the reference beam in a first position,
• c) measure a position of the reference beam signal captured by the light sensing system while the sample beam is switched off,
• d) switch on the sample beam and measure a fringe contrast of a signal captured by the light sensing system
• e) change by an increment the position of said at least one pivotally actionable direction change mirror (TM3, TM4),
• f) repeat steps c) to e) until the sum of increments corresponds to a pre-defined working range of the pivotally actionable direction change mirror (TM3, TM4),
• g) compare the fringe contrast measurements obtained for each increment and store in the look-up table (LUT) the position of the pivotally actionable direction change mirror (TM3, TM4) for the fringe contrast measurement with the highest value, in conjunction with the position of the rotating beam system;
• h) rotate by a small increment the rotating beam system and repeat steps b) to g) until the rotating beam system has completed a 360° rotation.