Laser scanning microscope

Abstract

Laser Scanning Microscope with an illumination beam path for illumination of a sample and a detection beam path for wavelength-dependent recording of the light from the sample, whereby filters for selection of the detection wavelengths are provided, characterized in that at least one graduated filter spatially variable in regard to the threshold wavelength between the transmission and reflection is provided in several partial beam paths for the selection of the wavelengths.

Description

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING

FIG. 1 is a schematic diagram of a prior art Laser Scanning Microscope;

FIG. 2 is a shaded sketch of a graduated filter;

FIG. 3 is a schematic diagram illustrating the principle of a beam path of a flexible NFT with the graduated filter of FIG. 2;

FIG. 4 is a graph illustrating different gradients of threshold wavelength of the graduated filter of FIG. 2; and

FIG. 5 is a schematic diagram showing displacement operation of the graduated filter.

DETAILED DESCRIPTION OF THE INVENTION

In describing preferred embodiments of the present invention illustrated in the drawings, specific terminology is employed for the sake of clarity. However, the invention is not intended to be limited to the specific terminology so selected, and it is to be understood that each specific element includes all technical equivalents that operate in a similar manner to accomplish a similar purpose.

As a flexible secondary beam splitter NFT, a so-called graduated filter (for example short-pass) is used. This is a filter, the threshold wavelengths for the transmission of which varies in dependence of the position of the filter, so that it behaves, for instance, like a short-pass filter with a threshold wavelength 500 nm at one place, while like a short-pass filter with a threshold wavelength of 600 nm at another place as shown in FIG. 2. In it, a graduated filter with continuous variation of the threshold wavelength between the reflective and the transmissive behavior is shown schematically.

FIG. 3 shows a sketch in principle of a beam path of a flexible NFT with a graduated filter. The collimated initial incident beam is split by a graduated filter NFT in the direction of two detection beam paths DE1 and DE2.

The graduated filter NFT is located in the direction of the detection behind the main dichroic beam splitter (MDB in FIG. 1), preferably in the beam path at the usual location for a secondary dichroic beam splitter SDS.

The graduated filter NFT is arranged in the detection beam path at an angle, usually at 45 degrees with respect to the optical axis of the incident beam and is movable along that angular position in the beam path in order to vary its optically effective threshold wavelength between transmission (in direction DE1) and reflection (in direction DE2).

Therefore, depending on the place where such a filter is illuminated, the transmission characteristic of this filter changes. A flexible NFT can thus be simply realized in that such a graduated filter is arranged displaceably in an otherwise fixed beam path. A sketch of the corresponding embodiment is shown in FIG. 2. The graduated filter, as shown in FIG. 5, is displaceable under motor control. The motor responds to a central actuation unit that may be part of a computer system.

Thus it is of advantage if the spectral splitting of the light does not take spatially in order to thereafter mirror the individual (arranged spatially separately) spectral parts in different directions, and instead of using several filters with fixed threshold wavelength, a graduated filter is used. Thereby the variation of the threshold wavelength of the filter above the cross section of the incident light beam should be smaller than the desired spectral resolution.

Following advantageous modifications of the invention form the subject matter of the present publication:

• • 1. It does not matter whether it involves a short pass or a long pass graduated filter. In one case, the shorter wavelengths go to detector DE1 and the longer ones to detector DE2, and in the other case, it is the other way round;
  • 2. A part of the filter can also be embodied as a glass plate without any filtering properties (the light passes through the NFT unobstructed, no deflection of light to the second branch) or as a mirror (complete deflection of light to the second branch);
  • 3. The filter can be designed with a linear (displaceable) form or, for instance, like a wheel (disk, rotatable);
  • 4. Instead of being continuous, the filter can be coated with a number of different filters in steps, or some parts of the filter can be in steps and the other parts can be continuous; and
  • 5. Band passes are also conceivable in place of the short pass or long pass characteristics.

Variation of the wavelength can be adapted according to the desired spectral resolution and need not necessarily be linear, concrete jumps in the threshold wavelengths are shown in FIG. 3.

Modifications and variations of the above-described embodiments of the present invention are possible, as appreciated by those skilled in the art in light of the above teachings. It is therefore to be understood that, within the scope of the appended claims and their equivalents, the invention may be practiced otherwise than as specifically described.

Claims

1. A laser scanning microscope for viewing a sample, the laser scanning microscope comprising: an optical axis:means for producing illumination rays along the optical axis for illuminating the sample;means for producing a detection beat path for wave-dependent recording of light from the sample;filter means for selecting the detection-wavelengths, wherein at least one graduated filter spatially variable in regard to the threshold wavelength between transmission and reflection is provided in the detection beam path for the selection of the detection-wavelengths.

2. The Laser Scanning Microscope according to claim 1, comprising means for disposing the graduated filter is a displaceable manner in the detection beam path so as to enable variation of the optically effective threshold wavelength of the graduated filter.

3. The Laser Scanning Microscope according to claim 1, comprising at least one additional partial beam path and at least one additional graduated filter arranged in the at least one additional partial beam path.

4. The Laser Scanning Microscope according to claim 2, further comprising a central actuation unit for controlling the displacement of the graduated filter.

5. The Laser Scanning Microscope of according to claim 4, wherein the displacement of the graduated filter is controlled by a motor receiving signals from the central actuation unit.

6. The Laser Scanning Microscope according to claim 4, wherein the central actuation unit provides for flexible assignment of the detection-wavelengths.

7. A beam splitter for use in a detection beam path of a Laser Scanning Microscope for wavelength-dependent splitting into a transmitted light and a reflected light, the beam splitter comprising: a graduated filter displaceable in the beam path for adjustable splitting the transmitted light and the reflected light.

8. The beam splitter according to claim 7, wherein the graduated filter comprises at least partially continuous spatial variation of the threshold wavelength between the reflected and the transmitted part of the detection light.

9. The beam splitter according to claim 7, wherein the graduated filter exhibits at least in part step-wise spatial variation of the threshold wavelength between the reflected and the transmitted part of the detection light.

10. The beam splitter according to claim 7, wherein the graduated filter is a short-pass filter, a long-pass filter or a band-pass filter.