Lumos Operation Manual Note WWW.MOTIC.COM MOTIC HONG KONG LIMITED If the equipment is used in a manner not specified by the manufacturer, the protection provided by the equipment may be impaired.
This instruction Manual has been prepared for users of the Motic “FS6” Epi-Fluorescence Attachment used in conjunction with the Motic PA53 Upright Microscope. For your own safety, read this manual carefully and thoroughly before using the product. Do not discard this manual. Always keep it near the product for easy reference. We are constantly endeavoring to improve our instruments and to adapt them to the requirements of modern research techniques and testing methods.
PRINCIPLE Fluorescence The phenomenon that occurs when light absorbed by a material, creates a molecular excitation that causes the material to re-emit energy in the form of light at a different wavelength. Fluorescence Microscope for Epi-Fluorescence The technique of fluorescence microscopy with Epi-illuminators is based on the adaptation ofthe vertical illuminator used in reflected light microscopy.
CONTENTS Principle 2 Notes for Safety/Safety Precautions 6 Warning Additional Safety Precautions Lumos Led/HBO100 Fluorescence Lamps 8 1. Nomenclature 11 1.1 Intended use of the Microscope 11 1.2 PA53 Bio 11 1.3 PA53 FS6 with Lumos LED Fluorescence Lamp 13 1.4 PA53 FS6 with HBO100 (Mercury lamp) 14 1.5 EpiFS6 Fluorescence Turret 15 1.6 Lumos, LED Fluorescence Lamp 15 1.7 Lumos Control Pad 16 1.8 HBO100 Mercury Lamp 17 1.9 MXH-100 Power supply unit 17 2.
3.5 3.6 Observation Tube 23 3.5.1 Adjusting the Interpupillar Distance 23 3.5.2 Adjusting the Diopter 23 3.5.3 Selecting the Light Path of the Trinocular Tube 24 Condenser 24 3.6.1 Centering the Condenser 24 3.6.2 Setting the Aperture Iris Diaphragm 26 3.7 Installing the Epi-Fluorescence Turret 27 3.8 Attaching the Fluorescence Illuminator 27 3.9 Installing the slider with the filter holder 29 3.10 Shutter Slider 29 3.11 Installing the Filter Cubes 29 3.11.
4.4 Using an oil immersion objective 47 4.5 Fluorescence Photomicrography 48 4.6 Video Fluorescence Microscopy 48 5. Troubleshooting Table 49 5.1 Electrical 49 5.2 Optical 49 6. Care And Maintenance 51 6.1 Do not disassemble 51 6.2 Lenses and filters 51 6.3 Cleaning or painted or plastic components 51 6.4 Disinfecting the Microscope 51 6.5 When not in use 51 6.6 For MXH-100 Power supply unit 52 7. Specification 53 7.1 Electrical Specification 53 7.
NOTES FOR SAFETY/SAFETY PRECAUTIONS To avoid being electrocuted, do not touch the lamp socket while the light is on as a high voltage current is being supplied to the bulb. Because the bulb produces a large amount of heat, do not touch the housing, socket or bulb when on. This instruction manual needs to be consulted in all cases where this symbol is used, in order to find out the nature of the potential HAZARD and any actions which have to be taken.
Because the bulb produces a large amount of heat, do not touch the housing, socket or bulb When it on or still hot. Make sure that the switch of voltage range is set to matches your available line voltage. Disconnect the power supply before replacement of fuse. To avoid the risk, make sure the MXH-100 starter is used with Motic lamp house.
WARNING ADDITIONAL SAFETY PRECAUTIONS LUMOS LED/ HBO100 MERCURY LAMP Ɣ While LED/HBO100 are generally safe illumination systems, precautions should still be taken with this powerful Fluorescence Illumination products. Ɣ When operating these products, please observe the following safety precautions at all times. Failure to do so may result in injuries or damage. Ɣ Strong UV light may be emitted from the products depending on the wavelength selected. Avoid eye and skin exposure at all times.
Ɣ For changing the bulb of the light source mercury lamp the usage of suitable protection equipment (Solid gloves and Face protection mask) is mandatory Ɣ WARNING: UV emitted from this product. Avoid eye and skin exposure to unshielded product. Ɣ WARNING: Possibly hazardous optical radiation emitted from this product. Do not look at operating lamp. Eye injury may result. Ɣ CAUTION: IR emitted from this product. Avoid eye exposure. Use appropriate shielding or eye protection.
Instrument Disposal Please observe the following safety notes for the disposal of the microscope: Defective Instruments, accessories and consumables should be disposed in compliance with the provisions of the local law. Instrument warranty The Instrument should only be used for microcopy applications mentioned and instructed in this operation manual. Please note the following information on the instruments warranty.
1. NOMENCLATURE 1.1 Intended use of the Microscope This Microscope is an optical instrument that has been designed to be used to observe and document magnified images of specimens in teaching, routine and research applications. Do not use this instrument for any other purpose than its intended use. 1.2 PA53 Bio (Fig.
(Fig.
1.3 PA53 FS6 with Lumos LED Fluorescence Lamp (Fig.
1.4 PA53 FS6 with HBO100 (Mercury lamp) (Fig.
1.5 EpiFS6 Fluorescence Turret (Fig. 5) 1.6 Lumos, LED Fluorescence Lamp (Fig.
1.7 Lumos Control Pad Control Pad (Fig.
1.8 HBO100 Mercury Lamp (Fig. 8) 1.9 MXH-100 Power supply unit (Fig.
2. SETTING UP THE INSTRUMENT Ɣ Avoid placing the instrument in areas exposed to dust, vibration, high temperature and high humidity and where it is difficult to unplug the power supply cord. Ɣ Avoid locations exposed to direct sunlight, under room lights. Ɣ Select location which allows easy access to power cord from products AC inletin the event of an emergency. 2.
3. ASSEMBLY INSTRUCTION 3.1 PA53 Microscope Follow the instructions provided in the microscope manual. For the purpose of performing simultaneous observation with phase contrast, attach the appropriate phase contrast components referring to the specific instructions provided. WARNING: Installation of upper part should be done by service engineer, not by customer!!! 3.2 Base 3.2.1 Voltage Indication and Switch Ɣ Push the brightness adjustment knob to switch transmitted / reflected light illumination.
3.2.3 Turn ON/OFF Intelligent Light function Ɣ Push IL button to toggle intelligent light function on and off. Ɣ The IL indicator LED 1 turns on when the intelligent light function turns on. Ɣ If the intelligent light function turns on, the brightness (lamp voltage) automatically changed preset value when the revolving nosepiece is changed. Ɣ To set new preset brightness, press IL button long while IL indicator LED turns on. (Fig. 12) 3.2.
3.3 Focusing Block 3.3.1 Replacing the Fine Adjustment Knob Ɣ The fine adjustment knob is designed detachable to prevent interference with hand during manipulation of the fine adjustment knob or XY-axis knob. Ɣ Usually attach the knob on the opposite side to the XY-axis knob. Ɣ Using the Allen screwdriver, loosen the clamping screw and remove the fine adjustment knob. (Fig. 14) 3.3.
3.3.3 Pre-focusing Lever Ɣ The pre-focusing lever ensures that the objectives does not come in contact with the specimen and simplifies focusing. Ɣ After focusing on the specimen with the coarse adjustment knob, turn this lever clockwise and lock. The upper limit on coarse adjustment movement is set at the locked position.
3.5 Observation Tube 3.5.1 Adjusting the Interpupillar Distance Ɣ While looking through the eyepieces, adjust for binocular vision until the left and right fields of view coincide completely. Ɣ The Index dot indicates the interpupillary distance. (Fig. 18) 3.5.2 Adjusting the Diopter Ɣ The diopter adjustment ring is used when the eyepiece has no function of diopter adjustment.
3.5.3 Selecting the Light Path of the Trinocular Tube Ɣ Slide the light path selector knob to select the desired light path. Light Path Selector Knob Indication Light Path Ratio Pushed-in position Binocular 100% Pulled-out position TV & Photo 100% (Fig. 20) 3.6 Condenser 3.6.1 Centering the Condenser Ɣ Turn the condenser height adjustment knob (Fig. 21) to raise the condenser to its upper limit. Ɣ Focus on the specimen using the 10x objectives. Ɣ Rotate the field iris diaphragm ring (Fig.
NOTE: The field iris diaphragm restricts the diameter of the beam of light entering the objectives and thus excludes extraneous light, improving image contrast. The diameter of the field iris should be adjusted for objective power to the extent that it just circumscribes the field of view. (Fig. 21) (Fig. 22) Fig.
3.6.2 Setting the Aperture Iris Diaphragm Ɣ The aperture iris diaphragm determines the numerical aperture of the illumination system. It has an effect of adjusting image resolution and contrast. Ɣ Since the contrast of microscope specimens is ordinarily low, setting the condenser aperture iris diaphragm to between 70 and 80% of the NA of the objectives in use is usually recommended.
3.7 Installing the Epi-Fluorescence Turret Please consult the illustrations provided while assembling the attachment Before Starting: Turn off the microscope power switch and unplug the power cord 3.8 Attaching the Fluorescence Illuminator Ɣ Fully loosen the four neck cover clamping screws (Fig. 26-3) of the microscope frame with provided 2.5mm Allen wrench. Remove neck cover (Fig. 26-4). Ɣ Place the Fluorescenceilluminator on the mounting surface. (Fig. 26-5) There are two position pins (Fig.
(Fig. 26-3) (Fig. 26-4) (Fig. 26-5) (Fig. 26-6) (Fig. 26-7) (Fig.
3.9 Installing the slider with the filter holder Ɣ Insert the slider into thefilterslider slot of the fluorescence attachment. (Fig. 27) (Fig. 27) 3.10 Shutter Slider (Fig. 28-1) Ɣ Blocking the optical path of the Epi-fluorescence by sliding the shutter to the left. (Fig. 28-2) (Fig. 28-1) (Fig. 28-2) 3.
Ɣ If the filter turret is not filled with five filter cubes, there is the risk that UV rays will reach the eyes through an empty space,causing serious harm. Ɣ Never perform lamp centering without the UV filter cube in the optical path as harmful UV radiation from the lamp may enter the eyes, possibly resulting in loss of vision.
(Fig. 29-3) (Fig. 29-4) (Fig. 29-5) (Fig.
3.11.2 Disassemble the filter set into the turret Step 1: Open the two clamp screw mounted Filter cube cover with the screwdriver delivered with Epi FS6 Turret package. (Fig. 30-1) Step 2: Lift head from plastic rivet (both sides of filter cube) (Fig. 30-2) Filter cube can be lifted up and removed (Fig. 30-2) Step 3: Replace the filter cube cover and secure with 2 clamp screws. (Fig. 30-3) Update filter ID Tag according to equipment (Fig. 30-1) (Fig. 30-2) (Fig.
3.12 Filter cube ID tags Ɣ Place the Filter cube ID tag on the front face of the Epi-fluorescence attachment.(Fig. 31-1) Ɣ Select the ID tags which represent the excitation method cubes in the turret and drop them into correspondingposition. (Fig. 31-2) (Fig. 31-1) (Fig. 31-2) 3.
3.14 Attaching UV light protective shield Ɣ Loosen UV light protective shield Clamp screws under Epi-fluorescence attachment body. (Fig.33-1) Ɣ Insert the UV light protective Shield – Slots are in the front of shield. (Fig. 33-2) Ɣ Secure in place with the clamp screws. (Fig. 33-3) (Fig. 33-1) (Fig. 33-2) (Fig. 33-3) 3.15 Mounting UV light blocking tube Ɣ Mount UV light blocking tube on the microscope condenser carrier and secure in place with condenser clamp screw. (Fig. 34) (Fig.
3.16 Attaching the Lumos lamp house Ɣ Attach the Mercury lamp house or Lumos LED lamp to the microscope using the dovetail on the Epi-fluorescence attachment. Ɣ Align the positioning pin on the Mercury lamp house (or Lumos) flange with the positioning groove on the dovetail mount of the fluorescence attachment (Fig. 35-1 / 35-3) Ɣ Secure the dovetail with the provided Allen Key as shown in Fig. 35-2 / 35-4. (Fig. 35-1) (Fig. 35-2) (Fig. 35-3) (Fig. 35-4) 3.16.
(Fig. 36) 3.16.2 Setting the reflected light intensity manager Ɣ It is effective when the PA53 FS6 revolving nosepiece with sensor and the LED lamp housing Lumos are used. Ɣ The desired reflected fluorescent brightness can be set per objective. Once the brightness for the objectives is set, selecting an objective adjusts the Lumos brightness automatically. 1 Switch to reflected fluorescent lighting path, (so that the light intensity indicator on the front of frame is lit in blue).
4 After 3 seconds, the microscope automatically memorizes the optimum brightness. 5 Set the optimum brightness of the reflected light path for the mounted objectives by repeating the above for each of them. 3.16.3 Setting the fluorescence turret channel manager Ɣ It is effective when the PA53 FS6 revolving fluorescence turret with sensor and the LED lamp housing Lumos are used.
3.17 Installing the lamp (Only for Mercury Lamp) NOTICE: Please follow all NOTES FOR SAFETY / SAFETY PRECAUTIONS and user personal protection equipment all the time Ɣ In order to prevent electric shock please always make sure to turn the power switch off, and unplug the power cord before installing or replacing the lamp. Ɣ Loosen the Mercury lamp house cover clamp screw (Fig. 38-1 -- Loosen) and remove the cover andthelamp socket assembly. (Fig. 38-2) (Fig. 38-1) Ɣ (Fig.
Ɣ Insert the lamp into the lower flexible lamp clamp base and tighten the clamp screw. (Fig. 38-5) Ɣ Insert the lamp into the upper lamp socket hole supported by the metal plate with the coolingfins and fasten the clamp screw. (Fig. 38-5) Ɣ Insert the bottom tab of lamp house cover over the inner metal surround of the lamp houseand pivot the lamp socket cover to close (Fig. 38-6). Secure with the clamp screw. (Fig.
3.18 Connecting the **Mercury lamp house to the power supply: * Please check the MXH-100 power supply instruction manual for more information. Turn OFF the power switch on the power supply. MXH-100 Power Supply (Fig.
3.19 Mecury Lamp House** * Please check the HBO100 Mecury Lamp instruction manual for more information. Lamp Housing (Fig.
3.20 Attaching the Lamp housing WARNING The bulb, the lamp socket and areas near these will be extremely hot during and right after use. Set the main switch off; disconnect the power cord from the wall outlet, then wait until lamp housing has cooled down before removing the Lamp housing. Ɣ Loosen the clamping screw (Fig. 41-1.1) with provided Allen screwdriver. Ɣ Mount the lamp housing to the connection port (Fig. 41-1.2) on the rear of the illuminator and press the lamphousing as far as it goes.
3.22 Changing the size of the aperture diaphragm (Fig. 42-2) Ɣ The aperture diaphragm determines the illuminated in the back focal plane of the microscope. Ɣ Pull or Push aperture diaphragm adjustment lever to change the size of the field aperture diaphragm. For normal observation, just the aperture diaphragm Ɣ Closing the aperture diaphragm will reduce the resolution but enlarge the depth of focus of the microscope (Fig. 42-1) (Fig. 42-2) 3.
4. MICROSCOPY Epi-Fluorescence Micoscopy 4.1 Selecting fluorescence filters: For best results, select excitation and emission filters with center wavelengths as close to the absorption and emission peaks as possible. Note: The center wavelength is situated at the midpoint of the bandwidth. It is not necessarily the peak transmission wavelength although with a symmetrical band the center wavelength and the peak wavelength are equal. 4.
4.3.1 Longpass Longpass filters only allow wavelengths above a certain Wavelength to pass through and will prevent light of lower wavelength from passing through. Longpass filters should be used when the application requires maximum emission and spectral differentiation is not necessary. This is generally the case for fluorophores that generate a single emitting species in specimens with reasonably low levels of background auto fluorescence.
Selecting fluorescence filters requires a thorough understanding of filter technology. This will enable the user to utilize stain and illumination selection to improve the image quality of the desired fluorescence signal. Selection of filter combinations also requires knowledge of the excitation and emission spectra of the stain.
4.4 Using an oil immersion objective Oil immersion objectives are labelled with the additional engraving “Oil” and are to be immersed in oil between the specimen and the front of the objective. The immersion oil supplied by Motic is synthetic, non-fluorescing and non-resining oil, with a refractive index of 1.515 Normally, a cover glass must be used with oil immersion objectives with a few exceptions.
4.5 Fluorescence Photomicrography For the basic procedure and key points of photomicrography, see the manuals provided with the photomicrographic equipment Since the specimen colour may fade effort must be made on minimizing the exposure of the specimen to irradiation both before, and during exposure. Select the area of interest without using fluorescence, use phase contrast or interference contrast.
5. TROUBLESHOOTING TABLE As you use your microscope and the Epi-Fluorescence attachment, you may occasionally experience a problem. The troubleshooting table below contains the majority of frequently encountered problems and the possible causes. 5.
Greasy residue on eyelens Room light is too bright Revolving nosepiece not clicked into position.
6. CARE AND MAINTENANCE 6.1 Do not disassemble Ɣ Disassembly may significantly affect the performance of the instrument, and may result in electric shock or injury and will void the terms of the warranty. Ɣ Never attempt to dismantle any parts other than described in this manual. If you notice any malfunction, contact your nearest Motic representative. 6.2 Lenses and filters Ɣ To clean lens surfaces or filters, first remove dust using an air blower.
Note: Ɣ If equipment is used in a manner not specified by the manufacturer, the warranty may be void. Ɣ To avoid getting wet, do not use the microscope near water. Proper handling of the microscope will ensure years of trouble-free service. If repair becomes necessary, please contact your Motic agency or our Technical Service directly. 6.6 For MXH-100 Power supply unit Ɣ Ensure the fan on the back panel of the power supply is kept free of obstructions.
7. SPECIFICATION 7.1 Electrical Specification Ɣ PA53 FS6: INPUT: AC100-240V~, 1.8-1A, 47~63Hz REFLECTED LAMP: 100W Halogen Ɣ PA53 FS6 Intermediate: INPUT: AC100-240V~, 3.2A, 47~63Hz OUTPUT: DC 5V/0.5A LAMP: 10W UV LED and 20W Blue, Green LED Ɣ PA53 Bio: INPUT: AC100-240V~, 1.8-1A, 47~63Hz REFLECTED LAMP: 100W Halogen 7.
7.4 Bulb replacement The lamp and the lamp house become very hot during and after a period of operation. Risk of burn – Do not touch the lamp during or immediately after period of operation. Make sure the lamp has cooled sufficiently before attempting to replace the lamp. Ɣ In order to prevent electric shock always turn the power switch off and unplug the power cord before installing or replacing the bulb. 7.
7.6 Miscellaneous Ɣ Automatic voltage selection device is applicable to worldwide voltage configuration, but it is recommended to use the power cable matches with the rated voltage of your area. Incorrect use of power cable may cause fire, or damage to the device Ɣ For external power cable, power cable with ground protection must be used. Ɣ To prevent electric shock, make sure the power switch is turned off before connecting the power cable.
7.
HPlan SAPO 4X/0.13, W.D=17.3mm, CG=0.17 HPlan SAPO 10X/0.3, W.D=11.7mm, CG=0.17 HPlan SAPO Series HPlan SAPO 20X/0.5, W.D=2.2mm, CG=0.17 Not recommend for Professional Flour Application HPlan SAPO 40X/0.75, W.D=0.7mm, CG=0.17 HPlan SAPO 100X/1.3 oil, W.D=0.1mm, CG=0.17 Plan UC PH 10X/0.25, W.D=17.4mm, CG=0.17, Ph1 Plan UC Phase Contract Series Plan UC PH 20X/0.45, W.D=0.8mm, CG=0.17, Ph2 Plan UC PH 40X/0.65, W.D=0.6mm, CG=0.17, Ph2 Plan UC PH 100X/1.25 Oil, W.D=0.16mm, CG=0.
FLUORESCENCE FILTER CUBES (Ø25mm Series) The fluorescence vertical illuminator can accommodate three filter blocks and a DIA-Filter dummy block (devoid of filters) that enables normal brightfield observation. Filter blocks are mounted on a slider inside the Epi-fluorescence attachment and can be inserted into the optical path by means of the excitation method change over knob.
STANDARD FLUORESCENCE FILTER SETS AT39002 EGFP/FITC/Cy2/AlexaFluor 488 Exciter AT480/30x Dichroic AT505DC Emitter AT39004 AT535/40m TRITC/Cy3/TagRFP/AlexaFluor 546 Exciter AT540/25x Dichroic AT565DC Emitter AT39010 AT605/55m Texas Red/mCherry/AlexaFluor 594 Exciter AT560/40x (EX) AT600DC (BS) Emitter AT635/60m (EM) AT39000 DAPI/Hoechst/AlexaFluor 350 Exciter :AT375/28x Dichroic :AT415DC Emitter:AT460/50m The emission at these wavelengths will be better detected by camera than by the unaided eye.
ARRANGEMENT OF FILTERS IN A FLUORESCENCE CUBE Upright Microscope It is often the case that specific combinations of excitation filter, emission filter and dichroic mirror areneeded to visualize and/or quantitate the fluorescence emission from a particular fluorescent species.
NOMENCLATURE Excitation Filter - EX A filter used in fluorescence microscopy designed to pass only those wavelengths, which excite fluorescence. Excitation Filter D350/50x The center wavelength is at 350nm; full bandwidth is 50nm [= ±25nm]. In some cases when the bandwidth is not specified, the letter "x" is used to define the filter as an exciter filter and is generally used for narrow band UV excitation filters, e.g. D350x.
DEFINITIONS OF FILTER TERMINOLOGY Bandpass Filters Bandpass filters transmit a band of wavelengths and block all light above and below the specified transmission range. These filters are characterized with respect to optical performance by their centre wavelength (CWL) and bandwidth, also referred to as the full width at half of maximum transmission (FWHM).
GG Green Glass – Longpass absorption glass from Schott Glassworks with cut-on wavelengths in the violet and blue-green regions HQ A designation for high-performance filters with narrow cuton and cutoff wavelengths LP Longpass Filters Longpass filters transmit a wide spectral band of long wavelength radiation while blocking short wavelength radiation. (See Figure Below) Narrowband Filters Narrowband filters with a very narrow band, typically 1 to 3nm wide.
SP Shortpass Filters Shortpass filters transmit a wide spectral band of short wavelength radiation and block long wave radiation. (See Figure Below) S/N Signal to Noise Ratio Ratio of intensity of signal to that of the background. Stokes shift The distance between the peak absorption and emission of a dye, usually in nm. Filters for isolating the wavelength of illumination: Shortpass and longpass filters, block or transmit wavelengths at specific cut-off wavelengths.
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