Microscope problem ~ ~ Please be knowledgeable, please ~

BIOX.CN 2005-4-1613: 38: 00 Source: Life Jingwei

Ordinary optical microscope is a precise optical instrument. In the past, the simplest microscope consisted of only a few lenses, but now the microscope used consists of a group of lenses. Ordinary optical microscope can usually magnify an object 1500-2000 times.

(A) the structure of the microscope

The structure of ordinary optical microscope can be divided into two parts: one is mechanical device, and the other is optical system. Only when these two parts are well coordinated can the microscope play its role.

1, mechanical device of microscope

The mechanical device of microscope includes lens holder, lens barrel, objective lens converter, stage, push rod, coarse screw, fine screw and other parts.

(1) The mirror mount is the basic support of the microscope, and consists of two parts: the base and the mirror arm. There is an object stage and the lens barrel connected with it, which is the basis for installing all parts of the optical amplification system.

(2) The lens barrel is connected with the eyepiece at the top and the converter at the bottom, and a darkroom is formed between the eyepiece and the objective lens (installed below the converter).

The distance from the rear edge of the objective lens to the rear end of the lens barrel is called the mechanical lens barrel length. Because the magnification of the objective lens is for a certain lens barrel length. With the change of lens barrel length, not only the magnification changes, but also the imaging quality will be affected. Therefore, when using a microscope, the length of the lens barrel cannot be changed at will. The international standard lens barrel length of microscope is 160mm, and this number is marked on the shell of objective lens.

(3) objective lens converter 3-4 objective lenses can be installed on the objective lens converter, usually three objective lenses (low power, high power, oil mirror). Nikon microscope is equipped with four objective lenses. By rotating the converter, any objective lens can be connected to the lens barrel as required, and form an amplification system with the eyepiece on the lens barrel.

(4) There is a hole in the center of the stage for light to pass through. The stage is equipped with a spring specimen holder and a pusher, which are used to fix or move the position of the specimen so that the object under the microscope is just in the center of the field of view.

(5) The pusher is a mechanical device for moving samples, which is made of a metal frame and has two pushing shafts, one horizontal and the other vertical. A good microscope is engraved with scales on the vertical and horizontal frames to form a very accurate plane coordinate system. If you need to observe a certain part of the specimen repeatedly, you can write down the values of the vertical and horizontal rulers during the first inspection, and then move the push rod according to the values to find the original specimen position.

(6) Thick Screw A thick screw is a machine that moves the lens barrel to adjust the distance between the objective lens and the specimen. The thick screw of the old microscope is screwed forward, and the lens descends close to the specimen. When testing a newly produced microscope (such as Nikon microscope), twist the right hand forward to make the stage rise and make the specimen close to the objective lens, otherwise, it will fall and the specimen will be separated from the objective lens.

(7) Micro screw The coarse screw can only roughly adjust the focal length. To get the clearest image, it needs to be further adjusted with the micro screw. The micro screw moves the lens barrel by 0. 1 mm (100 micron) per revolution. The thick and thin screws of the newly produced advanced microscope are * * * axes.

2. Microscope optical system

The optical system of microscope consists of reflector, condenser, objective lens and eyepiece. The optical system magnifies the object and forms an enlarged image of the object. See figure 1-2.

(1) Mirror Early ordinary optical microscopes used natural light to inspect objects, and a mirror was installed on the mirror base. The reflector consists of a plane and another concave mirror, which can reflect the light projected on it to the center of the condenser and illuminate the specimen. Concave mirror is used when the condenser is not used, and concave mirror can concentrate light. When a condenser lens is used, a flat mirror is usually used. The newly produced advanced microscope base is equipped with light source and current adjusting screw, and the light intensity can be adjusted by adjusting the current.

(2) The condenser is under the stage and consists of a condenser lens, an iris and lifting screws. The condenser can be divided into bright field condenser and dark field condenser. Ordinary optical microscopes are equipped with bright-field condenser, including Abbe condenser, Qi Ming condenser and dithering condenser. Abbe condenser will show excellent aberration and spherical aberration when the numerical aperture of the objective lens is higher than 0.6. Qi Ming condenser has a high degree of correction for chromatic aberration, spherical aberration and coma aberration, and is the best condenser in bright-field microscope, but it is not suitable for objective lens less than 4 times. Shake out the condenser can shake out the lens on the condenser from the optical path to meet the needs of large field of view illumination of low magnification objective lens (4 times).

The condenser is installed under the stage, and its function is to focus the light reflected by the light source on the sample through the reflector, so as to obtain the strongest illumination and make the image bright and clear. The height of the condenser can be adjusted so that the focus falls on the detected object to obtain the maximum brightness. Generally, the focus of a condenser is above 1.25mm, and its rising limit is below the stage plane by 0.1mm.. Therefore, it is required that the thickness of the slide should be between 0.8- 1.2 mm, otherwise the sample to be inspected will not be in focus, which will affect the microscopic inspection effect. There is also an iris diaphragm in front of the front lens group of the condenser lens, which can be enlarged and reduced, affecting the resolution and contrast of imaging. If the variable aperture is too large and exceeds the numerical aperture of the objective lens, a light spot will be produced. If the contracted iris aperture is too small, the resolution will decrease and the contrast will increase. Therefore, when observing, the field stop (microscope with field stop) is opened to the tangent of the periphery of the field of view by adjusting the aperture of the aperture, so that the objects not in the field of view are not irradiated by any light, thus avoiding the interference of scattered light.

(3) The objective lens installed on the front conversion mirror of the lens barrel makes the measured object be imaged by light for the first time, and the imaging quality of the objective lens has a decisive influence on the resolution. The performance of an objective lens depends on its numerical aperture (NA). The numerical aperture of each objective lens is marked on the housing of the objective lens. The larger the numerical aperture, the better the performance of the objective lens.

There are many kinds of objective lenses, which can be classified from different angles:

According to the different media between the front lens of the objective lens and the detected object, it can be divided into:

① The objective lens of the drying system takes air as the medium, such as the commonly used objective lens below 40×, and the numerical aperture is less than 1.

(2) Oil-immersed objective lenses often use aromatic asphalt as the medium, also known as oil lenses. The magnification is 90×— 100×, and the numerical aperture value is greater than 1.

According to the magnification of the objective lens, it can be divided into:

(1) low power objective lens refers to 1×-6×, and the NA value is 0.04-0.15;

② Dynamic objective refers to 6×-25×, and the NA value is 0.15-0.40;

(3) The high power objective lens is 25×-63×, and the NA value is 0.35-0.95;

④ The oil-immersed objective lens is 90×- 100×, and the NA value is 1.25- 1.40.

According to the aberration correction degree of the objective lens, it can be classified into:

① Achromatic objective lens is the most commonly used objective lens, and the shell is marked with the word "Ach", which can remove the chromatic aberration formed by red light and blue light. Microscope is usually used with Huygens eyepiece.

② Apo is marked on the objective shell of apochromatic objective lens, which can correct not only the chromatic aberration of red, blue and green light, but also the phase difference caused by yellow light. It is usually used with the compensation eyepiece.

(3) The special objective lens is based on the above objective lens, in order to achieve a certain observation effect. Such as: objective lens with correction ring, objective lens with field diaphragm, phase difference objective lens, fluorescent objective lens, strain-free objective lens, maskless objective lens, long working distance objective lens, etc. At present, the commonly used objective lenses in research include semi-apochromatic objective lens (FL), flat-field objective lens (Plan), flat-field apochromatic objective lens (PLAN) and super-flat-field objective lens (PLAN).

(4) Eyepiece The function of the eyepiece is to enlarge the real image amplified by the objective lens again and reflect the object image to the eyes of the observer. The structure of eyepiece is simpler than that of objective lens. The eyepiece of an ordinary optical microscope usually consists of two lenses, the upper lens is called "eyepiece" and the lower lens is called "field lens". An annular diaphragm made of metal or called "field diaphragm" is installed between the upper and lower lenses or under the two lenses. The intermediate image magnified by the objective lens falls on the plane of the field diaphragm, so an eyepiece micrometer can be placed on it.

Huygens eyepiece is often used in general optical microscope. For research purposes, eyepieces with better performance are generally selected, such as compensation eyepiece (K), flat field eyepiece (P) and wide field eyepiece (WF). Choose the photographic eyepiece (NFK) when taking pictures.

(2) Imaging principle of optical microscope

The magnification of the microscope is completed by the lens, and there are aberrations in Dan Toujing imaging, which affect the image quality. The lens group composed of Dan Toujing is equivalent to a convex lens with better magnification. Figure 1-4 is the imaging principle mode of microscope. AB is a specimen.

(3) the performance of the microscope

The resolution of microscope depends on various conditions of optical system. The observed object must be clear at high magnification. Whether an object can present a clear and fine structure after enlargement depends first on the performance of the objective lens, and then on the performance of the eyepiece and condenser.

1, the numerical aperture is also called aperture ratio (or aperture ratio), abbreviated as N.A. Both the objective lens and the condenser are marked with its numerical aperture. Numerical aperture is the main parameter of objective lens and condenser, and it is also the most important index to judge their performance. The numerical aperture is closely related to the performance of the microscope, which is directly proportional to the resolution of the microscope, inversely proportional to the focal depth and directly proportional to the square root of the mirror brightness.

The numerical aperture can be expressed by the following formula:

Noun (abbreviation of noun) A=n.sin α

2

Among them:

Dielectric diffraction rate between objective lens and sample

α-Mirror angle of objective lens

The so-called mirror angle refers to the angle between the light emitted by the object point on the optical axis of the objective lens and the edge of the effective diameter of the front lens of the objective lens, as shown in figure 1-5.

The rearview mirror angle α is always less than 180. Because the refractive index of air is 1, the numerical aperture of the drying objective is always less than 1, generally 0.05-0.95; If the oil-immersed objective lens is impregnated with fragrant tar (refractive index is 1.5 15), the maximum numerical aperture can be close to 1.5. Although the limit of numerical aperture is equal to the refractive index of immersion medium in theory, it is practically impossible to reach this limit from the perspective of lens manufacturing technology. Generally, the maximum numerical aperture of an advanced oil-immersed objective lens is 1.4.

The refractive indices of several substances are as follows:

The air is 1.0, the water is 1.33, the glass is 1.5, the glycerin is 1.47, and the fragrant asphalt is 1.52.

See figure 1-6 for the influence of refractive index of medium on the optical path of objective lens.

Step 2 solve

D can be expressed by the following formula:

D=λ/2N.A。

The wavelength of visible light is 0.4-0.7 micron, and the average wavelength is 0.55 micron. If an objective lens with a numerical aperture of 0.65 is used, D=0.55 micron /2×0.65=0.42 micron. This means that the detected object can be observed above 0.42 micron, but not below 0.42 micron. If an objective lens with a numerical aperture of 1.25 is used, D=2.20 microns. Any object longer than this value can be seen. Therefore, the smaller the D value, the higher the resolution and the clearer the image. According to the above formula, the wavelength can be reduced (1); (2) increase the refractive index; (3) Increase the angle of the mirror and improve the resolution. Microscopes and electron microscopes that use ultraviolet light as light source use short waves to improve the resolution to examine smaller objects. The resolution of the objective lens is closely related to the clarity of the image. The eyepiece does not have this performance. The eyepiece only magnifies the image produced by the objective lens.

3. Magnification:

When a microscope magnifies an object, it first magnifies the image through the objective lens, and the eyepiece magnifies the image at an apparent distance for the second time. Magnification is the ratio of the volume of the final image to the volume of the original object. Therefore, the magnification (v) of the microscope is equal to the product of the objective magnification (V 1) and the eyepiece magnification (V2), namely:

V=V 1×V2

A more accurate calculation method can be obtained from the following formula.

M= △ × D

F 1 F2

F 1= focal length of objective lens, F2= focal length of eyepiece△ = length of light pipe, and D= sight distance (=250 mm).

△ = Magnification of objective D = Magnification of eyepiece M= Magnification of microscope

F 1 F2

Let △ =160mm2 =150 = 4mm d = 250mm2 =150mm.

Then m = △× d =160× 250 = 40×16.7 = 668 times.

F 1 F2 4 15

4. Focus depth:

When observing a specimen under a microscope, the object image is clearest when the focus is on an image plane, which is the target plane. In the field of vision, in addition to the target plane, you can also see fuzzy objects above and below the target plane. The distance between these two planes is called focal depth. The focal depth of the objective lens is inversely proportional to the numerical aperture and magnification: that is, the greater the numerical aperture and magnification, the smaller the focal depth. Therefore, it is more careful to adjust the oil mirror than to lower the power mirror, otherwise it is easy for the object to slip and not be found.

Second, the use of microscope operation and matters needing attention

The microscope has a precise structure, so it must be used with care, and the following operation steps should be followed.

(1) Preparation before observation

1. When taking out the microscope from the microscope cabinet or mirror box, hold the mirror arm tightly with your right hand and the mirror base with your left hand, and transport the microscope to the experimental platform smoothly.

2. Put the microscope in front of your body on the left, about 10cm away from the edge of the table, and put a notebook or drawing paper on the right.

3. In the absence of a light source, the dimming microscope can use light or natural light to dim the light through the mirror, but it cannot use direct sunlight, which will affect the clarity of the object image and stimulate the eyes.

Turn the 10× objective lens into the aperture, open the iris aperture on the condenser to the maximum position, observe the brightness of the visual field in the eyepiece with the left eye, and turn the reflector to make the illumination of the visual field the brightest and most uniform. Use a flat reflector when the light is strong, and a concave reflector when the light is weak. The microscope with its own light source can adjust the light intensity by adjusting the current knob.

4. When observing the microscope by adjusting the optical axis center, the optical axes of the light source, condenser, objective lens and eyepiece in its optical system and the center of the diaphragm must be in a straight line with the optical axis of the microscope. For a microscope with a field stop, first narrow the field stop and observe it with a 10× objective lens. In the field of view, you can see the outline image of the spherical polygon of the field of view diaphragm. If this image is not in the center of the field of view, you can adjust it to the center with the two adjusting knobs on the outside of the condenser, and then slowly open the field of view diaphragm, so that you can see that the light beam spreads evenly around the field of view until the outline image of the field of view diaphragm is completely connected with the edge of the field of view, indicating that the light is coaxial.

(2) To observe and inspect any specimen with a low-power mirror, we must first get into the habit of observing with a low-power mirror. Because the low-power lens has a large field of view, it is easy to find the target and determine the location of the inspection.

Put the specimen on the stage, clamp it with the specimen, move the pusher so that the observed specimen is directly below the objective lens, turn the coarse adjustment knob to make the objective lens close to the specimen, observe with the eyepiece, slowly raise the lens barrel (or lower the stage) with the coarse adjustment knob until an object image appears, and then use the fine adjustment knob to make the object image clear. Move the specimen with a pusher, find a suitable target image, and move it to the center of the field of view for observation.

(3) High-power lens observation The high-power objective lens is converted on the basis of observation by the low-power objective lens. In a good microscope, the low-power and high-power lenses are in the same focus. In general, the conversion of high-power objective should not touch the slide or the cover glass on it. If different types of objective lenses are used, please observe from the side when replacing the objective lens to avoid collision between the lens and the slide. Then observe from the eyepiece, adjust the illuminance to make the brightness moderate, slowly adjust the coarse adjustment knob to make the stage rise (or the lens barrel fall) until the object image appears, then adjust it with the fine adjustment knob until the object image is clear, find the part to be observed, and move it to the center of the field of view for observation.

(4) The working distance of the oil-immersed objective lens (the distance between the front lens surface of the oil-immersed objective lens and the object to be inspected) is very short, generally within 0.2 mm. In addition, the oil-immersed objective lens of the general optical microscope has no "spring device". Therefore, special care should be taken when using the oil-immersed objective lens to avoid crushing the specimen and damaging the objective lens due to careless focusing.

Use the oil mirror to do the following:

1. First, raise (or lower) the lens barrel by about 2 cm with the coarse adjustment knob, and turn out the high-power lens.

2. Drop a drop of aromatic asphalt on the microscopic part of the slide specimen.

3. From the side, slowly raise the stage (or lower the lens barrel) with the coarse adjustment knob, so that the oil-immersed objective lens is immersed in the aromatic asphalt, and the lens is almost in contact with the specimen.

4. Observe from the eyepiece, enlarge the field stop and the iridescent stop on the condenser (the oil mirror with field stop opens the large field stop), and adjust the condenser to fully illuminate the light. Slowly lower the stage (or raise the lens barrel) with the coarse adjustment knob, and when the object flashes, adjust it to the clearest with the fine adjustment knob. If the oil mirror has left the oil surface and can't see the object, you must observe it from the side again and repeat the above operation.

5. After the observation, put down the stage and dig out the oil mirror. First, wipe off the oil stain on the lens with mirror paper, then dip the mirror paper in a little mixture of ether and alcohol (2 parts of ether and 3 parts of pure alcohol) or xylene to wipe off the residual oil on the lens, and finally wipe it with mirror paper for 2-3 times (be careful to wipe it in one direction).

6. Restore all components, rotate the objective lens converter so that the objective lens is not opposite to the light hole of the stage, but in an open position, then lower the lens barrel to the minimum, put down the condenser, and cover the eyepiece with a clean handkerchief to prevent the objective lens from being contaminated with dust. Finally, clean the mechanical parts such as the stage with soft gauze, and then put the microscope back into the cabinet or mirror box.

A metallographic microscope suitable for observing and photographing metallographic structures in the visible spectrum range at room temperature.

Project \ Parameters and Performance \ Grade Primary Metallographic Microscope, Intermediate Metallographic Microscope and Advanced Microscope

Performance bright field, (photography) bright field, dark field, polarization, photography, (binocular) (phase contrast) (microhardness) bright field, dark field, polarization, photography, binocular, projection, automatic exposure, (phase contrast) (microhardness), (interference) (interference phase contrast)

basis

basis

let go

big

The speed of things

Kagami Akira field 10X, 40X, (10X) 4X, 10X, 25X, 40X, 63X, 100X 2.5X, 4x,10x, 25X, 40X, 63X.

Dark field-10X, 25X 10X, 25X, 40X

Phase contrast-10X, 40X 10X, 40X, 63X

Eyepiece 5X, 10X, 12.5X 5X, 10X, 12.5X, 16X 5X, 10X, 12.5X,/kloc-0.

Objective Achromatic Flat-field Achromatic Flat-field Achromatic or Flat-field Semi-apochromatic Flat-field Apochromatic

Filter yellow, green, blue yellow, green, blue yellow, green, blue.

Observation form vision monocular and binocular

Total photographic magnification-100 times, 200 times, 500 times, 1000 times, 200 times, 400 times, 500 times, 800 times and 1000 times.

Width -8.25× 12 cm2 12× 16.5 cm2

Projection diameter-not less than φ 250mm

The workbench can move vertically and horizontally, and can rotate 360. The minimum reading is 1. The longitudinal and transverse moving range is not less than 10mm, and it can rotate 360, with a minimum reading of 6. The vertical and horizontal moving range shall not be less than 15mm.

The fine focus range of the fine focus mechanism is not less than 1.8mm, and the dividing value is 0.002mm.

Accessories must include-10X reticle eyepiece, 0.0 1mm micrometer, granularity plate 120 or 135 camera, automatic exposure device, 10X reticle eyepiece, 0.0 1mm micrometer and granularity.

Purchase 100X achromatic objective lens, photographic device, 10X reticle eyepiece, 0.0 1mm micron binocular eyepiece, phase contrast device, microhardness device, microhardness device and interference phase contrast device.

It is used for general metallographic analysis in factory site and school laboratory, metallographic analysis and research in factories and universities, as well as large and medium-sized factories, universities and scientific research departments.