JP3258349B2 - Epi-illumination fluorescence microscope - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an epi-illumination type fluorescence microscope having an optical element used for switching an excitation wavelength.

[0002]

2. Description of the Related Art For example, an apparatus for switching an excitation wavelength in an epi-illumination type fluorescence microscope is provided with a plurality of filter units (two in the drawing) as shown in FIG. Each of these filter units includes a support member having a moving mechanism for moving to the observation optical path, an optical substrate attached to the support member, and only a wavelength necessary for exciting fluorescence formed on the optical substrate. And an excitation filter. The switching of the excitation wavelength for irradiating the sample is performed by sliding the filter unit into the observation optical path as shown by the arrow.

[0003]

As described above, each filter unit is constituted by an independent optical substrate and a supporting member for supporting the optical substrate. For this reason, thickness errors and variations in parallelism occur between the optical substrates. Due to the thickness error and the variation of the parallelism, if the operation of switching the excitation wavelength is performed at the time of sample observation, the refraction state at the filter will be different, and the optical path toward the observation system will not exactly match before and after the switching operation. I will.

Further, if there is a processing error in the substrate support member, the moving mechanism, or the like in each filter unit, the filter unit will not be reproduced at the same position before and after the switching operation of the filter unit, and the illumination system for irradiating the sample will be described. The light from is blurred.

As described above, optical misalignment occurs due to errors in the thickness of the optical substrate, variations in parallelism, processing errors in the substrate support member, and the like. Is shifted. This hinders various uses, such as when taking a fluorescence photograph such as multiple excitation and multiple exposure.

SUMMARY OF THE INVENTION The present invention has been made in view of the above problems, and provides an epi-illumination type fluorescence microscope which does not cause optical misalignment before and after the switching operation of a wavelength or the like in an optical element. The purpose is to.

[0007]

In order to solve the above-mentioned problems, an epi-illumination type fluorescence microscope according to the present invention comprises: an illumination system for emitting light of a plurality of wavelengths necessary for exciting fluorescence; A single optical substrate provided with a plurality of types of optical functional films on the same surface for extracting only light having a wavelength necessary to excite a specific fluorescent light from a plurality of types of wavelengths emitted from the illumination system, respectively. While holding the optical substrate on the same surface, a movable member for switching an optical functional film of the optical substrate disposed in the observation optical path, and a movable member formed on the microscope main body, guiding the movable member, and A guide mechanism for switching the optical function film into the optical path, and an observation system for observing the sample based on fluorescence from the sample excited by light extracted by the optical function film of the optical substrate, And

[0008]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An epi-illumination type fluorescence microscope according to an embodiment of the present invention will be specifically described below with reference to the accompanying drawings. FIG. 1 is a diagram schematically showing an optical path in an epi-illumination fluorescence microscope.
In this figure, an optical element 10 is placed in a portion 1 surrounded by a square.
Is arranged.

The light emitted from the illumination system 5 is
Only the wavelengths that are reflected by 0 and necessary to excite the fluorescence are extracted and illuminate the specimen 9 via the objective lens 7. The reflected light, fluorescence, and the like from the specimen 9 pass through the optical element 10 again through the objective lens 7 and are guided to the observation system.

The structure of the optical element 10 will be specifically described with reference to FIG. FIGS. 2A and 2B are a plan view and a side view, respectively, of the optical substrate 11 on which the optical functional film 13 (in this embodiment, dichroic filters 13a and 13b having different spectral characteristics) is applied. . The formation of the optical functional film 13 on the optical substrate 11 is performed by a normal thin film forming technique, for example, vacuum evaporation, sputtering, or the like. The optical substrate 11 on which the optical function film 13 is formed is held by a movable mechanism member 20 described below.

FIG. 2C is a plan view showing the structure of the movable mechanism member 20, and FIG.
It is sectional drawing along the line. The movable mechanism member 20 has a support surface 21 inclined at 45 ° with respect to the optical axis.
The optical substrate 11 on which the optical function film 13 is formed is held on the support surface 21. Support surface 21
The dichroic filters 1 having different spectral characteristics.
Holes 23a and 23b are formed at positions corresponding to 3a and 13b, respectively, and guide the reflected light from the sample to the observation optical system. The movable mechanism member 20 has a hole 27 for guiding light from the illumination system to the dichroic filter, and a hole 29 for guiding light reflected by the dichroic filter to the sample. Of course, these holes 27 and 29 are formed corresponding to the hole 23a.

Further, the movable mechanism member 20 is formed with a switching sliding portion 25, which engages with a guide mechanism (not shown) provided on the microscope main body, and To slide. As a result, it is possible to switch the excitation wavelength by sliding the movable mechanism member 20.

As described above, since the dichroic filters 13a and 13b having different characteristics are formed on the same optical substrate 11, no optical misalignment occurs due to the thickness error of the optical substrate and the variation of the parallelism. . In this case, an influence based on a thickness error of the optically functional film can be considered, but since this film thickness is extremely thin, it can be ignored in practice. Further, since the single movable member 20 is configured to slide, no optical misalignment due to a processing error occurs.

In this embodiment, the number of optically functional films is two, but the number can be increased according to the wavelength required to excite fluorescence. Further, the mechanism for sliding the movable mechanism member 20 is not limited to the illustrated one, and may be configured by using, for example, a roller or a rack and pinion. Furthermore, in the present embodiment, a dichroic filter having different spectral characteristics is used as an optical functional film for fluorescence observation, but as an optical element of the present invention, a polarizing beam splitter, a half mirror, or the like is used depending on the optical device. A combination of those having characteristics may be used.

[0015]

As described above, according to the epi-illumination type fluorescence microscope of the present invention, when the switching operation of the wavelength and the optical path is performed by the optical element, the optical misalignment does not occur before and after the switching operation. Absent.

[Brief description of the drawings]

FIG. 1 is a diagram schematically showing an optical path in an epi-illumination type fluorescence microscope.

FIGS. 2A and 2B are a plan view and a side view showing an optical substrate on which dichroic filters having different spectral characteristics are adhered; FIG. 2C is a plan view showing a configuration of a movable mechanism member; D) is a cross-sectional view along the line DD in FIG.

FIG. 3 is a view schematically showing an apparatus for switching an excitation wavelength in a conventional epifluorescence microscope.

[Explanation of symbols]

10 optical element, 11 optical substrate, 13a, 13b dichroic filter (optically functional film), 20 movable member.

Continuation of the front page (58) Field surveyed (Int.Cl. ⁷ , DB name) G02B 21/00 G02B 5/00 G02B 26/00

Claims (1)

(57) [Claims] An illumination system that emits light of a plurality of wavelengths required to excite fluorescence, and a wavelength required to excite a specific fluorescence from the plurality of wavelengths emitted from the illumination system, respectively. A single optical substrate provided with a plurality of types of optically functional films on the same surface for extracting only light of the same type; holding the optical substrate on the same surface; A movable member for switching the functional film; a guide mechanism formed on the microscope main body for guiding the movable member to switch the optical functional film of the optical substrate into an optical path; and extracting by the optical functional film of the optical substrate. And an observation system for observing the sample based on fluorescence from the sample excited by the emitted light.