超声波细胞粉碎机的安装和操作
细胞破碎技术—超声波破碎
细胞破碎技术——超声波破碎法摘要：细胞破碎技术的基本概念及其基本方法，重点介绍了从超声波破碎仪及超声波破碎常见的问题与解决方法上介绍了超声波破碎法。
关键词：细胞破碎方法超声波破碎仪常见问题正文：一、细胞破碎阻力细菌——几乎所有细菌的细胞壁都是由肽聚糖（peptidoglycan）组成，它是难溶性的聚糖链（glycan chain），借助短肽交联而成的网状结构，包围在细胞周围，使细胞具有一定的形状和强度。
短肽一般由四或五个胺基酸组成，如L-丙氨醯-D-谷氨醯-L-赖氨醯-D-丙氨酸。
而且短肽中常有D-胺基酸与二氨基庚二酸存在。
破碎细菌的主要阻力是来自于肽聚糖的网状结构，其网结构的致密程度和强度取决于聚糖链上所存在的肽键的数量和其交联的程度，如果交联程度大，则网结构就致密。
酵母菌——酵母细胞壁的最里层是由葡聚糖的细纤维组成，它构成了细胞壁的刚性骨架，使细胞具有一定的形状，覆盖在细纤维上面的是一层糖蛋白，最外层是甘露聚糖，由1,6一磷酸二酯键共价连接，形成网状结构。
在该层的内部，有甘露聚糖-酶的复合物，它可以共价连接到网状结构上，也可以不连接。
与细菌细胞壁一样，破碎酵母细胞壁的阻力主要决定于壁结构交联的紧密程度和它的厚度。
真菌——霉菌的细胞壁主要存在三种聚合物，葡聚糖（主要以β-1,3糖苷键连接，某些以β-1,6糖苷键连接），几丁质（以微纤维状态存在）以及糖蛋白。
最外层是α-和β-葡聚糖的混合物，第2层是糖蛋白的网状结构，葡聚糖与糖蛋白结合起来，第3层主要是蛋白质，最内层主要是几丁质，几丁质的微纤维嵌入蛋白质结构中。
与酵母和细菌的细胞壁一样，真菌细胞壁的强度和聚合物的网状结构有关，不仅如此，它还含有几丁质或纤维素的纤维状结构，所以强度有所提高。
植物细胞——对于已生长结束的植物细胞壁可分为初生壁和次生壁两部分。
初生壁是细胞生长期形成的。
次生壁是细胞停止生长后，在初生壁内部形成的结构。
目前，较流行的初生细胞壁结构是由Lampert等人提出的“经纬”模型，依据这一模型，纤维素的微纤丝以平行于细胞壁平面的方向一层一层敷着在上面，同一层次上的微纤丝平行排列，而不同层次上则排列方向不同，互成一定角度，形成独立的网路，构成了细胞壁的“经”，模型中的“纬”是结构蛋白（富含羟脯氨酸的蛋白），它由细胞质分泌，垂直于细胞壁平面排列，并由异二酪氨酸交联成结构蛋白网，径向的微纤丝网和纬向的结构蛋白网之间又相互交联，构成更复杂的网路系统。
超声机操作规程
一、目的：为了使各操作人员按规范作业，保证安全，提高生产效率，特制订本规程。
二、适用范围：适用于超声波的操作。
三、操作程序：1、按照标示接上适当的电源（220-250V），若电压不稳定需附装稳压器，同时确保电线、开关制完好无破裂、漏电等，并接上安全的地线。
2、接通机器电源及压缩空气，气压设定4Kg/cm。
3、进行操作时，先打开电源总制，接通气压，以空机检查各个按制是否正常，尤其应注意“紧急停止制”以免在生产中出现事故。
4、锁紧上模，“音波检查”拨到“手动”，同时按下机台两边绿色按钮，使模具下降，通过调整机头高度及焊头行程螺母，使上模与下模相距1mm。
5、按下机台中间红色钮，使机头上升。
音波检查钮拨回到自动。
6、调整主机右侧的微调旋钮，边调边按动主机上的音波检查钮，使振幅指示最小。
7、与上模相对锁紧下模，调整延迟时间，使上模在接触待熔接物前，即发振。
8、在下模上放入待熔接的产品，试熔接，若上下不重合，则调整下模，直至两边重合，若熔接不够，则增加熔接时间，反之，减少熔接时间，调试正确后即可进入正常作业。
四、注意事项1、没受过培训，没有超声波上岗证的员工，禁止进行操作。
2、严禁两个人同时操作同一台超声波。
3、在开机时，严禁将手放在超声模上，防止开机时由于故障，焊头自行下行伤人。
4、在超声产品过程中，禁止用异物塞住其中一个拎手及用单手操作，避免受伤。
5、调整时，音波发振时间不能超过10S。
6、上，下模不能相碰，以免伤到模具。
7、音波发振不停时，应立即切断电源。
8、先发振再熔接。
6、在维修保养过程中，应关掉电源和气压，在超声波静止状态下进行，若需电源时应做好绝缘，防火等措施。
7、在按下双拎手过程中，若发现胶件没放好或其它紧急需停下来现象，要立刻按下超声机中间红色按扭，使超声机停止工作，以免损伤超声波模或伤人，若不幸被伤，应立即送往医务室医治。
超声波破碎仪使用说明书
USER’S GUIDEAUTOTUNE SERIESHIGH INTENSITY ULTRASONIC PROCESSORMICROPROCESSOR CONTROLLED100 Watt Model • 130 Watt ModelTABLE OF CONTENTSWarrantyImportant SafeguardsLow Surface Tension Liquids - Organic SolventsSECTION 1 – INSTALLATIONInspectionElectrical RequirementsInstalling the Ultrasonic ProcessorSECTION II – OPERATIONPrinciples of Ultrasonic DisruptionFunctions of Controls, Indicators, and ConnectorsPreparations for UseUsing the Ultrasonic ProcessorSECTION III – SERVICE INFORMATIONOverload ConditionReturn of EquipmentSECTION IV – OPERATING SUGGESTIONS AND TECHNIQUESThe Ultrasonic Processor supplied with this instruction manual is constructed of the finest material and the workmanship meets the highest standards. It has been thoroughly tested and inspected before leaving the factory and when used in accordance with the procedures outlined in this manual, will provide you with many years of safe and dependable service.Rev. April 2004IMPORTANT SAFEGUARDSREAD BEFORE INSTALLING ORUSING THE EQUIPMENTYour Ultrasonic Processor has been designed with safety in mind. However, no design can completely protect against improper usage, which may result in bodily injury and/or property damage. For your protection and equipment safeguard, observe the following warnings at all times, read the operating instructions carefully before operating the equipment, and retain this instruction manual for future reference. If the ultrasonic Processor is used in a manner contrary to that specified in this instruction manual, the protection features designed into the unit may be impaired.• When mounting the probe, always clamp the converter housing. Never clamp the probe.• Make sure the Ultrasonic Processor is properly grounded via a 3-prong outlet.• High voltage is present in the power supply. Do not remove the cover. Refer all servicing to qualified service personnel.• To avoid electric shock, disconnect the electrical power cord before removing the cover prior to servicing.• Never operate the power supply unless it is connected to the converter.• Never secure anything to the probe, except at the nodal point (point of no activity).• Never touch a vibrating probe.• Never allow a microtip to vibrate in air for more than 10 seconds.• Never operate a threaded probe without a replaceable tip.• Air-cool the converter when sample temperature exceeds 100° C, and when working at high intensity for more than 30 minutes.• It is recommended that a sound abating enclosure or ear protection be used when operating the Ultrasonic Processor.CAUTIONLOW SURFACE TENSION LIQUIDS – ORGANIC SOLVENTS The probes (solid or with a replaceable tip) are tuned elements that resonate at a specific frequency. When using a ½” (13 mm) probe with a replaceable tip, if the replaceable tip is removed or isolated from the rest of the probe, the probe will no longer resonate at that frequency, and the power supply will fail.Unlike aqueous (water based) solutions, which rarely cause problems, samples containing solvents or low surface tension liquids are problematic. These liquids penetrate the probe/replaceable tip interface, and force the particulates into the threaded section isolating the tip from the probe.When processing samples containing solvents or low surface tension liquids ALWAYS use a solid probeSECTION 1 – INSTALLATIONINSPECTIONPrior to installing the Ultrasonic Processor, perform a visual inspection to detect any evidence of damage, which might have occurred during shipment. Before disposing of any packaging material, check it carefully for small items.The Ultrasonic Processor was carefully packed and thoroughly inspected before leaving our factory. The carrier, upon acceptance of the shipment, assumed responsibility for its safe delivery. Claims for loss or damage sustained in transit must be submitted to the carrier.If damage has occurred, contact your carrier within 48 hours of the delivery date. DO NOT OPERATE DAMAGED EQUIPMENT. Retain all packing materials for future shipment.ELECTRICAL REQUIREMENTSThe Ultrasonic Processor requires a fused, single phrase 3-terminal grounding type electrical outlet capable of supplying 50/60 Hz at 100 volts, 115 volts, 220 volts, or 240 volts, depending on the voltage option selected. For power requirements, check the label on the back of the unit.Should it become necessary to convert the unit for different voltage operation, proceed as follows.1. Ensure that the power cord is not connected to the electrical outlet.2. Open the fuse holder cover using a small screwdriver.3. Pull out the red fuse holder from its housing.4. To convert from 100/115V to 220/240V replace the two 3 Amp slow blow fuses,with 1.6 Amp fuses.5. To convert from 220/240V to 100/115V reverse the procedure above.6. Rotate the fuse holder 180º from its original position, and reinsert it into itshousing. For 100/115V operation the voltage displayed should be 115. For220/240V operation the voltage display should be 220.7. Change the electrical power cord as required.The Ultrasonic Processor should be installed in an area that is free from excessive dust,dirt, explosive and corrosive fumes, and extremes of temperature and humidity.SECTION II – OPERATIONPRINCIPLES OF ULTRASONIC DISRUPTIONThe ultrasonic power supply converts 50/60 Hz line voltage to high frequency electrical energy. This high frequency electrical energy is transmitted to the piezoelectric transducer within the converter, where it is changed to mechanical vibrations. The vibrations from the converter are intensified by the probe, creating pressure waves in the liquid. This action forms millions of microscopic bubbles (cavities), which expand during the negative pressure excursion, and implode violently during the positive excursion. As the bubbles implode, they cause millions of shock waves and eddies to radiate outwardly from the site of collapse, as well as generate extremes in pressures and temperatures at the implosion sites. Although this phenomenon, known as cavitation, lasts but a few microseconds, and the amount of energy released by each individual bubble is minimal, the cumulative amount of energy generated is extremely high. The larger the probe tip, the larger the volume that can be processed but at a lesser intensity. For information regarding the processing capability of each probe, consult the tables below.FUNCTIONS OF KEYS, CONTROLS, INDICATORS, AND CONNECTORS (cont.)REAR PANEL9 pin D-sub connector Connects to external actuation device, and enable power and frequency monitoring.Footswitch jack Connects to the footswitch cable.Coax connector Connects to the converter.Power module Connects to the electrical line cord and encases the fuse(s).9-PIN D-SUB CONNECTORPin No. Description1 Notconnected2 Notconnected3 Notconnected4 Enables connection to a frequency counter.5 Enables connection to an external power monitor (5 mv = 1 watt)6 Ground7 Energizestheultrasonicswhen connected to ground.8 and 9 Enables the intensity to be remotely adjusted using an external 10k potentiometer. See belowPREPARATION FOR USE1. Plug the electrical line cord into the electrical outlet, if the unit is already on; depress2. If the optional footswitch is used, insert the plug into the jack located on the rearpanel. Make sure that the plug is inserted forcefully all the way in.3. If the converter / probe assembly is not already assembled; screw the probe into theconverter and secure with wrenches provided.4. Mount the converter / probe assembly in a laboratory stand, secure the clamp to theupper section of the converter housing only. Do not secure the clamp to the probe.5. Connect the converter cable to the power supply.USING THE ULTRASONIC PROCESSORand the following control parameters.AMPLITUDE: The amplitude is the only parameter that must be set in order for the Ultrasonic Processor to be operational. The other control parameters – Timeand Pulse, do not have to be set for continuous operation. AMPL. displays theamplitude selected e.g. 40%. To set the amplitude when the ultrasonics is off,press the AMPL key and the numeric keys for a 40% reading on the screen –Ampl 40%, and the ENTER/REVIEW key.The screen will display:The Ultrasonic Processor is now ready for continuous operation. To energize the ultrasonics, press the START key or the footswitch. To de-energize the ultrasonics, press the key or release the footswitch. If the Time or Pulse functions must be used, refer to the appropriate paragraph(s) below.To increase or decrease the amplitude in small increments when the ultrasonics is on, depress the AMPL key to display Amplitude Setting on the screen, then depress the ▲or ▼key, as required.Immerse the microtip half way into the sample. If the probe is immersed to an insufficient depth, air will be injected into the sample, causing the sample to foam.Since the amplitude required is application dependent and subject to the volume and composition of the sample, it is recommended that the amplitude be selected empirically, by increasing or decreasing the level of intensity as required as the sample is being processed.TIMER:In the pulsed mode the processing time will be different from theelapsed time because the processing time function monitors and controls only the ON portion of the duty cycle. For example, for 1 hour processing time, theelapsed time will be 2 hours if the ON and OFF cycle are set for 1 second. To set the processing time, press the TIMER key.The screen will display:Using the numeric keys, set the processing time as required:Press the ENTER/REVIEW key. The screen will display:PULSER:By inhibiting heat build-up in the sample, the pulse function enablessafe treatment of temperature sensitive samples at high intensity. In addition,pulsing enhances processing by allowing the material to settle back under theprobe after each burst. The ON and OFF pulse duration can be set independently from 01 second to 59 seconds. During the OFF portion of the cycle, the redindicator on the key will illuminate. If the OFF portion of the cycleexceeds three seconds, a cautionary message - Sonics in OFF Cycle - will warnthe operator against touching the ultrasonic probe. To set the pulser, press PULSE key.The screen will display:Using the numeric keys, set the ON portion of the cycle, and press the ENTER/REVIEW key.The screen will display:Using the numeric keys set the OFF portion of the cycle.The screen will display:Press the ENTER/REVIEW key.The screen will display:REVIEW: The REVIEW function provides a “window” on the process by displaying various operating parameters without process interruption. Pressing theENTER/REVIEW key repeatedly during processing will consecutively display the following information.a) Selected amplitude:e.g. Amplitude 40%b) Selected processing time and elapsed processing time:e.g. Set 0:30:00 Time 0:22:10c) Selected pulsing cycle, and actual pulsing cycle:e.g. Pulse 01 01 / 01 00d) Amount of power in watts, and accumulated amount of energy in JOULESdelivered to the probe:e.g. 20 watts 0000000 Joulese) Elapsed time since processing was initiated:e.g. Elapsed time 0:44:20SECTION III – SERVICE INFORMATIONYour Ultrasonic Processor was designed to provide you with years of safe anddependable service. Nevertheless, because of component failure or improper usage, the possibility does exist that it might not perform as it should, shut down due to an overload condition or that it will stop working all together. The most probable causes formalfunction are listed below and should be investigated.The unit was plugged into an electrical outlet that provides a different voltage fromthat required. See Electrical Requirements.The probe and/or microtip is not secured properly.If the probe has a replaceable tip, the tip is not secured properly, or the probe hasbeen used with samples containing solvents or low surface tension liquids.1. the electrical outlet.2. holder from its housing.3.Replace the fuse(s).4.disconnect the probe from the converter.5. reads above 10 watts, either the converter or power supply has failed and thecomplete Ultrasonic Processor should be returned for repair.OVERLOAD CONDITIONIf the Ultrasonic Processor stops working, and an OVERLOAD indication is displayed onRETURN OF EQUIPMENTIt is suggested that an Ultrasonic Processor in need of repair be sent back to the factory.In order to receive prompt service; always contact the factory before returning anyinstrument. Include date of purchase, model number and serial number. For instruments not covered by the 3-year warranty, a purchase order should be forwarded to avoidunnecessary delay. Care should be exercised to provide adequate packing to insureagainst possible damage in shipment. The Ultrasonic Processor should be sent to the “Service Department” with all transportation charges prepaid and return of shipment indicated.Please obtain a Return Authorization Number prior to returning the instrument.SECTION IV - OPERATING SUGGESTIONS AND TECHNIQUES DISRUPTING CELLSThe disruption of cells is an important stage in the isolation and preparation of intracellular products. From research levels through to production, many areas of biotechnology, particularly recombinant technology, necessitate the use of ultrasonics for cell disruption. Although some biological products are secreted from the cell or released during autolysis, many others require sonication to release intracellular material. Cell disruption focuses on obtaining the desired product from within the cell, and it is the cell wall that must be disrupted to allow cell contents extraction.Single-cell organisms (micro-organisms) consist of a semipermeable, tough, rigid outer cell wall surrounding the protoplasmic membrane and cytoplasm. The cytoplasm is made up of nucleic acid, protein, carbohydrates, lipids, enzymes, inorganic ions, vitamins, pigments, inclusion bodies, and about 80% water. In order to isolate and extract any of these substances from inside the cell, it is necessary to break the cell wall and protoplasmic membrane. In some cases the cell may excrete the desired substance without assistance, but in most cases, the cells must be lysed and sonicated in order for these substances to be released. Breaking cell membranes and releasing the contents present significant challenges. The process must be fast and thorough to maximize the protein yield. Because the energy applied must be great enough to break the cell membranes or walls, yet gentle enough to avoid physically or chemically damaging cell contents, the Vibra-Cell with its variable intensity capability is ideally suited for this application.The level of intensity that should be used is application dependent. For example high intensity might be recommended for the break up of cells, but should never be used when the release of intracellular components might be objectionable e.g. Organelle isolation. The ability to control the amplitude at the probe tip is a prerequisite for process optimization. And because each application requires its own set of processing parameters, due to variation in volume and composition, the optimum amplitude can only be determined empirically. When processing a new sample, it is recommended that the amplitude be set first at 50% (30% with a microtip) and then increased of decreased as required.Yeast, gram-positive bacteria, and to a lesser extent, gram-negative bacteria have considerably harder cell walls in comparison to animal cells, and require relatively high power for cell disruption.Gram negative bacteria typically require 10 to 15 minutes of processing, while staphylococcus requires 20 to 30 minutes.Microorganisms differ greatly in their sensitivity to ultrasonic disintegration. For example, the most readily disintegrated are the rod-like forms (bacilli), while the spherical organisms (cocci) are much more resistant. The group Mycobacteria, to which the tuberculosis organism belongs, is particularly difficult to disrupt. Generally, animal cells are more easily disintegrated that plant cells, and red blood cells are more readily disintegrated than muscle cells because they lack a protective cell wall.Cellular disruption is the first step in RNA isolation and one of the most critical steps affecting yield and quality of the isolated RNA. Typically, cell disruption needs to be fast and thorough. Slow disruption, for example placing cells or tissue in guanidinium isothiocyanate (GITC) lysis solution for a long time prior to sonication, may result in RNA degradation by endogenous RNases released internally. This is especially a concern when working with tissues high in endogenous RNase such as spleen and pancreas. Disrupting frozen tissue is more time consuming and cumbersome that processing fresh tissue, but freezing samples is sometimes necessary. Samples are usually frozen when, 1) they are collected over a period of time and thus, cannot be processed simultaneously; 2) there are many samples, 3) samples are collected in the field, or 4) mechanical processing of fresh samples is insufficient for thorough disruption. A mortar and pestle or bag and hammer are typically used when the starting material is frozen. RNA will remain intact in tissues for a day at 37ºC, a week at 25ºC a month at 4ºC and indefinitely at subzero temperatures.Ultrasonic processing will typically cause the temperature of the sample to increase especially with small volumes. Since high temperatures inhibit cavitation, the sample temperature should be kept as low as possible - preferably just above its freezing point. This can be accomplished by immersing the sample vessel in an ice-salt-water bath. Temperature elevation can also be minimized by using the pulser.Increasing hydrostatic pressure (typically 15-60 psi) and viscosity can enhance cell disruption. For microorganisms, the addition of glass beads in the 0.5 to 1mm size range promotes cell disruption. Beads are almost a prerequisite when working with spores and yeast. A good ratio is one volume of beads to two volumes of liquid. Glass beads are available from Cataphote, Inc. P.O. Box 2369, Jackson, Mississippi 39225-2369 USA, phone (800) 221-2574 or (601) 939-4612, FAX (601) 932-5339, Jayco Inc. 675 Rahway Ave., Union NJ 07083 USA, phone (908) 688-3600, FAX (908) 688-6060 or Sigmund Lindner GmbH. P.O. Box 29. D-95483 Warmensteinach, Germany. Phone (49) 0 92 77 9 94 10, FAX (49) 0 92 77 9 94 99.When processing difficult cells such as yeast, pretreatment with an enzyme is beneficial. Lysozyme, byaluronidase, glycosidase, glucalase, lyticase, zymolase and lysostaphin digestion are among the enzymatic methods frequently used with yeast and Lysozyme with bacteria. Enzymatic treatment is usually followed by sonication in a GITC lysis buffer. Collogenase may be used with collogen, lysostaphin with staphylococcus, and trypsin hyaluronidase with liver and kidney.If enzymes cannot be used, the following procedures should be considered: Freezing the sample at -70°C overnight, then thawing it in water immediately prior to ultrasonic processing.Most animal tissues can be processed fresh (unfrozen). It is important to keep fresh tissue cold and to process it quickly (within 30 minutes) after dissection. When working with fresh tissue, the cells must be sonicated immediately at the time the GITC lysis solution is added. This can be done by dispensing the lysing solution in the tube, adding the tissue and immediately sonicating. Samples should never be left sitting in lysis solution, undisrupted. Large samples of hard tissues should be first treated in a blender or a mechanical homogenizer.Animal tissues that have been frozen after collection should be disrupted by grinding in liquid nitrogen with a mortar and pestle. During this process, it is important that the equipment and tissue remain at cryogenic temperatures. The tissue should be dry and powdery after grinding. Grinding should be followed by thorough sonication in a GITC lysis buffer. Processing frozen tissue in this way is cumbersome and time consuming, but effective.Cultured cells are normally easy to disrupt. Cells grown in suspension are collected by centrifugation, rinsed with PBS to remove culture medium, and then lysed by sonicating in a GITC lysis buffer. Placement of the vessel on ice while washing and lysing the cells will further protect the RNA from endogenous RNases released during the disruption process.Soft, fresh plant tissue can often be disrupted by sonicating in a lysis buffer. Other plant tissues, like pine needles, need to be ground dry, without liquid nitrogen. Some hard, woody plant materials require freezing and grinding in liquid nitrogen prior to being ultrasonically processed. Plant cell suspension cultures and calluses can typically be sonicated in a lysis buffer within 2 minutes. The diversity of plants and plant tissue make it impossible to give a single recommendation for all. However, most plant tissues typically contain polysaccharides and polyphenols that can coprecipitate with RNA and inhibit downstream assays. Treating a plant tissue lysate with polyvinylpyrrolidone (PVP) will precipitate such problematic components from the lysate before the actual RNA isolation is carried out.Whenever possible, the tissues should be diced very small to permit movement within the liquid. Tough tissues such as skin and muscle should be macerated first in a blender or the like for about 10 seconds, and confined to a small vessel during ultrasonic treatment. If sub-cellular particles are desired intact, the amplitude should be kept low, and the processing time increased.Yeast can be extremely difficult to disrupt because their cell walls may form capsules or nearly indestructible spores. To process yeast, sonicate in a tube containing the sample, guanidinium-based lysis buffer and small glass beads (0.5 – 1 mm). Pretreatment withzymolase, glucalase and / or lyticase to produce spheroplasts that are readily lysed may also be useful.To disrupt filamentous fungi, scrape the mycelial mat into a cold mortar, add liquid nitrogen and grind to a fine powder with a pestle. The powder can then be thoroughly sonicated in lysis buffer to solubilize completely. As fungi may also be rich in polysaccharides, pretreatment with polyvinylpyrrolidone (PVP) may be beneficial. Bacteria, like plants, are extremely diverse; therefore, it is difficult to make one recommendation for all bacteria. Ultrasonic processing will lyse most Gram positive and Gram negative bacteria, including mycobacteria. Although it is recommended that glass beads and lysis solution be used; it is possible to lyse some Gram negative bacteria by sonicating in lysis solution without beads. Bacteria cell walls can be digested with lysozyme to form spheroplasts. Gram positive bacteria usually require more rigorous digestion and longer processing time. The spheroplasts are then lysed with sonication in GITC lysis buffer.Disruption of cells found in soil and sediments is accomplished one of two ways. One technique isolates the bacterial cells from the material prior to the RNA isolation procedure. This is accomplished by homogenization of wet soil in a mechanical blender followed by a slow speed centrifugation to remove fungal biomass and soil debris. The supernatant is centrifuged again at a higher speed to pellet the bacteria cells. Cells can then be lysed as described above for bacteria. Other techniques describe RNA isolation from the soil or sediment directly. For example, one method requires soil to be added to a diatomaceous earth and lysis buffer, and then sonicated. The sample is then centrifuged to remove solid debris.Always immerse the probe deep enough below the surface of the sample to inhibit aerosoling or foaming, foaming substantially reduces cavitation. Processing at a lower power setting without foam is much more effective than processing at a higher power setting with foam. Decreasing the power, increasing processing time and lowering the temperature of the sample will usually prevent aerosoling and foaming. Do not use any antifoaming agents or surfactants.During cavitation, free radicals are formed which, if they are allowed to accumulate, can greatly affect the biological integrity of the sample by reacting with proteins, polysaccharides, or nucleic acids. Although during short periods of processing their formation is not normally considered a problem; for longer durations, the addition of free radical scavengers such as, carbon dioxide, N2O, cysteine, reduced glutahione, dithiothreitol or other SH compounds, might be beneficial. Saturating the sample with a protective atmosphere of helium or nitrogen gas, or dropping a small pellet of dry ice in the sample, will also inhibit free radical formation. Whereas it is true that gas is required for effective cellular disruption, it is not necessary that the vapor phase be oxygen or air since any gas except carbon dioxide will work just as well.21Various methods can be used to measure the efficiency of the disruption. For example, a visual count can be made using a microscope.For greater accuracy, a protein assay could be used. This procedure is widely recognized as a good method for measuring cell disruption by taking into account the amount of protein released after disruption. The disrupted cells are then tested and checked against this number for percentage breakage.There are several types of protein assays. One commonly used is the Folin Reaction (Lowry Assay) method, as it is comparatively simple and provides consistent results. This colorimetric method has a sensitivity to protein of around 8 µg / mL in the assay solution.The assay turns blue in the presence of proteins due to the reaction of copper ions in the alkaline solution with protein and the reduction of phosphomolybdate- phosphotungstic acid in the Folin reagent by aromatic amino acids in the treated protein.Fractional protein release, Rp, is calculated using the following equation and multiplying the result by 100:Rp = Cf – CbCt – CbCf = Free proteinCt = total proteinCb = Background proteinThis gives the actual disruption percentage, taking into account the background levels of protein before disruption.Since the greatest concentration of energy is beneath the probe, it is imperative that the sample be kept as close to the tip as possible, liquids are easily processed because the free moving cells circulate repeatedly below the probe. Solid materials however have a tendency to be repelled by the ultrasonic, and should be processed in a vessel large enough to accommodate the probe, yet small enough to restrict sample movement. For small samples, conical shaped test tubes are recommended.Allowing the probe to contact the vessel will decrease the power output, and cause minute grey glass particles to migrate into the sample. Although these glass particles will not adversely affect the chemical composition of the sample, they will form a thin grey layer on centrifuging. If the probe has to come in contact with a solid sample, use a standard 20mm (3/4”) diameter stainless steel centrifuge tube cut to 70mm (3”) length. Do not use a glass tube. Microtips must never allowed to come in contact with anything but the liquid, because the stress resulting at the point of contact with a hard surface will cause the microtip to fracture. Although larger probes will not fracture if they come in contact with a glass vessel, they may cause the vessel to fracture.Before each application, place the tip in water or alcohol and energize the power supply for a few seconds to remove any residual substances. If concerned about contamination from previous use, clean the probe with a 20% Virkon solution and rinse with distilled water. Because Virkon®, Dupont's broad spectrum cleaner / disinfectant is effective against a wide range of virus, bacteria and fungi, it is ideally suited for that purpose. For further information regarding Virkon® contact BiosafetyUSA, Inc. 10220 NW 50thStreet, Sunrise Florida 33351, 954-578-6580 *124 or e-mail info@For critical application, probes may be autoclaved.22。
超声机的使用流程
超声机的使用流程1. 概述超声机是一种常用的医疗设备，用于进行超声检查和诊断。
本文档介绍了超声机的使用流程，包括准备工作、设备操作和注意事项，以帮助用户正确地操作超声机。
2. 准备工作在使用超声机之前，需要进行以下准备工作：•确保超声室环境整洁、安静，并保持适宜的温度和湿度。
•检查超声机的电源线是否正常连接，并保证电源供应稳定。
•准备好超声探头，并检查探头表面是否有损坏或污染。
•打开超声机主机，并进行系统自检，确保设备正常运行。
3. 设备操作超声机的操作步骤如下：3.1 设置扫描模式超声机提供多种扫描模式，包括B模式（二维超声）、M模式（运动超声）和Doppler模式（血流超声）等。
根据具体的检查需要，选择合适的扫描模式。
3.2 准备扫描区域根据需要进行超声检查的部位，将超声凝胶涂抹在该部位，并确保良好接触。
确保扫描区域干净、无障碍物。
3.3 调整超声参数根据检查需要，调整超声机的参数，包括增益、扫描深度、频率、对比度等。
确保超声图像清晰可见。
3.4 开始扫描将超声探头对准扫描区域，保持适当的角度和压力，开始进行扫描。
注意保持手部稳定，避免晃动。
3.5 观察和记录在扫描过程中，通过观察超声图像，注意机器显示的相关参数，如距离、速度等，记录需要的信息。
4. 注意事项在使用超声机时，需要注意以下事项：•避免将超声探头与针或其他尖锐物体接触，以防损坏探头。
•操作超声机时，注意手部的卫生，勤洗手、戴手套，以防交叉感染。
•使用过程中如发现超声机异常噪音、图像不清晰或其他问题，应立即停止使用，并联系专业人员进行检修。
•对于不同部位的超声检查，应使用适当的探头和参数，以获得准确的检查结果。
•在使用超声机时，应注意患者的隐私和舒适度，保持良好的沟通和配合。
5. 总结本文档介绍了超声机的使用流程，包括准备工作、设备操作和注意事项。
正确地操作超声机可以提高检查的准确性和效果，同时保障患者的安全和舒适度。
用户在使用超声机时，应遵循本文档所述的步骤和注意事项，并注意设备的维护和保养，以延长超声机的寿命和提高其性能。
超声波治疗仪的操作流程
超声波具有穿透性 强、能量易于集中 、作用效果显著等 特点。
适用范围与禁忌症
适用范围
适用于多种软组织损伤和炎症的治疗，如肌肉、韧带、筋膜等。
禁忌症
对于急性炎症、恶性肿瘤、出血倾向等患者不宜使用。
操作前准备
确认设备电源已连接并处于待 机状态。
检查患者身体状况，向患者解 释治疗过程及注意事项。
准备好适当的耦合剂，以确保 声波能够有效地传递到患者患
注意事项及安全提示
治疗前检查
选择合适的剂量
在使用超声波治疗仪前，应进行全面的检查 ，确保患者没有禁忌症，了解患者的病情和 需要。
根据患者的实际情况选择合适的剂量，以确 保安全有效的治疗。
注意个体差异
避免禁忌部位
每个人的身体状况和反应都有所不同，因此 需要根据个体差异调整治疗方案，以确保最 佳效果。
对于有创伤、溃疡、炎症或其他禁忌部位的 部位，应避免使用超声波激
如果在使用过程中出现皮肤刺激、 瘙痒、红疹等症状，应立即停止使 用，并进行相应的处理。
疼痛加剧
如果在治疗过程中出现疼痛加剧或 异常感觉，应立即停止使用，并进 行相应的处理。
过敏反应
如果在使用过程中出现过敏反应， 如呼吸急促、喘息等症状，应立即 停止使用，并进行相应的处理。
定期检查与校准
设备校准
按照生产厂家提供的校准方法 ，定期对设备进行校准，确保
设备性能稳定、准确。
部件更换
定期更换易损件，如探头、过 滤器等，保证设备正常运行。
仪器保养
定期对设备进行保养，如清洁 内部、润滑等，确保设备长期
稳定运行。
常见故障及排除方法
电源故障
仪器不启动
电源线破损、电源插座接触不良——更换 电源线、清理插座。
超微粉碎机微粉机使用说明
超微粉碎机微粉机使用说明一、产品介绍：超微粉碎机，又称为微粉机，是一种粉碎分级设备，主要用于对各种脆性材料进行超细粉碎。
该设备采用优质合金刀具，具有高效、精细的粉碎效果，广泛应用于化工、冶金、食品、医药等行业。
本使用说明将详细介绍超微粉碎机的使用方法和注意事项，以保证设备的正常使用。
二、使用方法：1.将超微粉碎机放置在坚固的平台上，接通电源。
2.根据粉碎物料的特性，选择相应的刀具进行装配，并将刀具安装到转子上。
3.打开送料斗，并将待粉碎物料均匀地加入到斗口中。
5.通过调节转速控制器，选择合适的转速，以达到所需的粉碎效果。
通常来说，转速越高，粉碎效果越细。
6.在设备运行过程中，可以适时地加入润滑液，以降低设备摩擦和磨损。
7.物料粉碎完成后，关闭设备，并等待设备停止运转后，再打开安全门进行取样。
三、注意事项：1.在使用超微粉碎机前，务必仔细阅读并理解本使用说明。
4.在给料过程中，应将待粉碎物料均匀地分布于转子刀具周围，以免过大或过小的物料导致粉碎不均匀。
5.在加料的过程中，应保持盘中液压的稳定性，避免因流速过小或过大导致设备堵塞或物料飞溅。
6.在启动设备前，应先检查设备周围是否有杂物或障碍物，并确保设备不受干扰。
7.在设备运行时，应定期检查刀具的磨损情况，如有磨损应及时更换。
8.在操作过程中，应保持设备清洁，并随时清理设备内部和外部的粉尘和杂质。
9.在使用润滑液时，应选择合适的润滑剂，并按照规定的用量和方法添加。
10.在清洁设备时，应先切断电源，并使用干净的抹布或刷子清理设备，切勿使用水直接清洗设备。
几种细胞粉碎的方法
几种细胞粉碎的方法细胞破碎的方法:一、机械破碎法：是指利用捣碎机、研磨器或匀浆器等将细胞破碎开来。
1. 高速组织捣碎：将材料配成稀糊状液，放置于筒内约1/3体积，盖紧筒盖，将调速器先拨至最慢处，开动开关后，逐步加速至所需速度。
此法适用动物内脏组织、植物肉质种子等。
2. 玻璃匀浆器匀浆：先将剪碎的组织置于管中，再套入研杆来回研磨，上下移动，即可将细胞研碎，此法细胞破碎程度比高速组织捣碎机为高，适用于量少和动物脏器组织。
二、物理破碎法：指利用温度差、压力差或超声波等将细胞破碎开来。
1：用一定功率的超声波处理细胞悬液，使细胞急剧震荡破裂(借助超声的震动力破碎细胞壁和细胞器）。
机制：可能与强声波作用溶液时，气泡产生、长大和破碎的空化现象有关，空化现象引起的冲击波和剪刀力使细胞裂解。
超声波破碎的效率取决于声频、声能、处理时间、细胞浓度和细胞类型等。
（使用时注意降温，防止过热）。
2. 高压破碎：细胞悬浮液从高压室的环状隙喷射到静止的撞击环上，被迫改变方向经出口管流出。
此过程中细胞经历了高速造成的剪切的碰撞及高压到常压的变化，从而破碎释放内含物。
这是一种温和的、彻底破碎细胞的较理想的方法。
3. 反复冻融法：将细胞在-20度以下冰冻，室温融解，反复几次，由于细胞内冰粒形成和剩余细胞液的盐浓度增高引起溶胀，使细胞结构破碎。
三、化学破碎法：指利用甲醛、丙酮等有机溶剂或表面活性剂作用于细胞膜，使细胞膜的结构遭到破坏或透性发生改变。
有些动物细胞，例如肿瘤细胞可采用十二烷基磺酸钠（SDS）、去氧胆酸钠等细胞膜破坏。
浓度一般为1mg/ml。
四、酶学破碎法：选用合适的酶，使细胞壁遭到破坏，进而在低渗溶液中将原生质体破碎开来。
细菌细胞壁较厚，可采用溶菌酶处理效果更好。
裂解液标准配方：50mM Tris-HCl(pH8.5~9.0), 2mM EDTA, 100mM NaCl, 0.5% Triton X-100, 1mg/ml溶菌酶。
超声波仪器使用方法
超声波仪器使用方法超声波仪器是一种常用于医学、工业和科学研究领域的设备，它可以通过高频声波的传播和反射来获取和分析材料的结构和性质。
本文将介绍超声波仪器的使用方法，帮助读者更好地了解和操作这一设备。
一、超声波仪器的基本原理超声波仪器利用声波在材料中的传播和反射来获取信息。
当超声波传播到不同密度和介质性质的材料中时，它们会发生折射和反射。
通过测量这些折射和反射，我们可以得到材料的结构、密度和缺陷等重要信息。
二、准备工作在开始使用超声波仪器之前，需要进行一些准备工作：1. 确保仪器处于正常工作状态，如电源是否连接，设备是否正常启动。
2. 检查仪器所需的探头是否安装正确，接触良好。
3. 对于需要进行声速校准的情况，要保证浸泡液的温度和浓度正确。
三、操作步骤1. 设置参数根据具体需要，设置合适的工作频率、脉冲宽度和接收增益等参数。
这些参数的选择应根据待测材料的特性和实验要求进行调整。
2. 放置探头将探头放置在待测材料的表面或浸没在液体中，确保探头与材料之间有足够的接触面积和良好的耦合。
3. 发送超声波信号通过触摸屏或按钮等设备，发送超声波信号至探头。
仪器会将信号发送至待测材料中，并记录各种声波的传播和反射信息。
4. 数据采集和分析仪器会实时显示从待测材料中接收到的声波信号，并对其进行处理和分析。
通过观察信号的特征和变化，可以判断材料的结构、密度和缺陷等信息。
5. 结果解读根据仪器显示的数据和分析结果，对材料进行评估和判断。
可以参考标准曲线、图像对比或经验知识等方法，进行结果解读和分析。
四、注意事项1. 使用超声波仪器时，需要保持仪器和探头的清洁和完好。
避免杂质或损坏影响信号的传播和接收。
2. 不同材料的声速、衰减等参数不同，需要根据具体情况进行校准和调整。
3. 在操作过程中，要注意安全。
避免探头与尖锐物体接触，防止仪器受损或人员受伤。
总结：超声波仪器是一种非常有用的设备，它能够通过声波的传播和反射来获取材料的结构和性质信息。
超声波设备使用说明书
超声波设备使用说明书一、产品简介超声波设备（以下简称“设备”）是一款专业的医疗设备，采用超声波技术，广泛应用于医疗诊断和治疗领域。
本设备具备高精度、便携、易操作等优点，可为医务人员提供准确、可靠的超声波成像和治疗功能。
二、安全注意事项1. 使用本设备前，请仔细阅读说明书并按照指南操作。
2. 本设备只适用于经过专业培训的医疗人员使用。
3. 请确保设备的电源和接线符合安全要求。
4. 设备操作期间，请确保周围环境安静，以保证诊断的准确性。
5. 不得自行拆卸设备或进行维修，如有故障，请联系售后服务部门。
三、设备操作步骤1. 准备工作a. 确保设备已连接电源并处于正常工作状态。
b. 擦拭设备外壳，确保设备表面无污垢和损坏。
c. 检查并连接适当的探头，确保其固定稳定。
2. 启动设备a. 按下电源开关，设备开始启动。
b. 在设备界面上选择相应的操作模式。
3. 超声波成像操作a. 将液体凝胶涂抹于患者体表与探头之间。
b. 将探头轻轻贴附于患者体表，并保持适当的角度和压力。
c. 调整设备参数，如增益、深度等，以获得清晰的图像。
d. 根据医疗需要，在所得图像上进行测量、标记等操作。
4. 超声波治疗操作a. 根据医疗需求选择相应的治疗模式。
b. 在操作区域涂抹适量的导入剂。
c. 将治疗探头轻轻贴附于患者体表，并调整适当的治疗参数。
d. 进行治疗，注意观察患者的反应和治疗效果。
e. 治疗结束后，关闭设备并将探头清洁、消毒。
四、操作注意事项1. 操作过程中，请注意设备和探头的温度，避免烫伤患者或自身。
2. 操作过程中请确保患者舒适，并与患者保持沟通以了解患者的感受。
3. 严禁在敏感部位、破损皮肤或感染部位使用本设备。
4. 操作过程中请注意设备的保养和维修，避免影响设备使用寿命。
五、设备保养1. 每次使用后，请将设备的探头进行彻底清洁和消毒。
2. 定期检查设备的电源线和连接线是否正常，如有损坏请及时更换。
3. 定期进行设备的校准和维护，确保设备工作的准确性和可靠性。
超声波细胞粉碎机原理
超声波细胞粉碎机原理简介超声波细胞粉碎机是一种利用强超声在液体中产生空化效应，对物质进行超声处理的多功能、多用途的仪器，能用于动植物组织、细胞、细菌、芽胞菌种的破碎，同时可用来乳化、分离、分散、匀化、提取、脱气、清洗及加速化学反应等等。
该机广泛应用于生物化学、微生物学、药物化学、表面化学、物理学、动物学、农学、医学、制药等领域教学、科研、生产。
生物产业的发展，应用超声波细胞粉碎机所做的实验要求也随之提高，如对样品温度的测定、控制，低温冷却样品及整机的智能化程度的提高等等，都提出了新的要求。
基本原理超声波细胞粉碎机是利用超声波在液体中的分散效应，使液体产生空化的作用，从而使液体中的固体颗粒或细胞组织破碎．超声波细胞粉碎机由超声波发生器和换能器二大部分组成。
超声波发生器将市电转变成18-21KHz交变电能供给换能器。
锆钛酸钡压电振子是换能器的心脏，它随交变电压以18-21KHz频率作伸缩弹性形变，换能器随之作纵向机械振动。
振动波通过浸入在生物溶液中的钛合金变幅杆产生空化效应，激发介质里的生物微粒剧烈振动。
特点1、自动调频起振，无需手工调节。
2、有温度保护探头，可有效防止在破碎过程中由于温度升高破坏样品。
3、可根据温度探头反馈的信号，通过“常开/常闭”开关驱动外部温度控制单元的工作。
4、标配的探头在1/2”以下可通过MICROTIP向下转换，实现微量样品的破碎。
5、有密闭杯（CUP）及连续流（FILLCELL）等不同样式的探头供选购。
6、有96孔酶标板破碎头及双针探头（保证实验平行性）探头供选购。
7、跟踪显示输出实际有效功率。
8、有九个程序记忆功能，方便用户编程及调用。
9、可调整“工作/间歇”比例时间循环脉冲式工作，时间范围从0.5秒～1。
小时，提高破碎效果，有效防止温升。
10、有保护微型探针的提问菜单，可以有效防止误操作损坏机器。
用途超声波粉碎机能用于粉碎动、植物细胞、细菌、牙孢或组织。
分散稀土和各类无机矿物粉。
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超声波细胞粉碎机的安装和操作
(1)、装置地：反应釜应安装在符合防爆要求的高压操作室内，在装备多台反应釜时，应分
开放置，每两台之间应用安全的防爆墙隔开，每间操作室均应有通向室外的通道和出口，当
存在易爆介质时应保证设备地通风良好。
(2)、打开包装后检查超声波细胞粉碎机有无损坏，根据设备型号按结构图将设备安装起来，
所配备件按照装箱单查清。加热方式如果是导热油电加热，请按照使用温度购买相应型号的
导热油(注意：导热油绝对不允许含有水分)加入，加入时将夹套上部的加油口打开并将夹套
中上部的油位口打开，通过加油口往里加油待油位口流油时即可，后将油位口拧死，勿将加
油口拧死以免产生压力。
(3)、釜体、釜盖的安装及密封：釜体和釜盖采用垫片或锥面与圆弧面的线接触，通过拧紧
主螺母使它们相互压紧达到良好的密封效果，拧紧螺母时必须对角对称多次逐步加力拧紧，
用力均匀，不允许釜盖向一边倾斜，以达到良好的密封效果，在拧紧主螺母时不得超过规定
的拧紧力矩40~120N.M范围，以防密封面被挤坏或超负荷磨损，密封面应特别加以爱护，每
次安装之前用比较柔软的纸或布将上下密封面擦拭干净，特别注意不要将釜体、釜盖密封面
碰上疤痕，若合理操作可使用上万次以上，密封面破坏后，需重新加工修复方可达到良好的
密封性能，拆卸釜盖时应将釜盖上下缓慢抬起，防止釜体与釜盖之间的密封面相互碰撞。如
果密封是采用垫片密封(四氟、铝垫、铜垫、石棉垫等)，通过拧紧主螺母便能达到良好的密
封效果。
4.阀门、压力表、安全阀的安装通过拧紧正反螺母，即达到密封的效果，联接两头的圆弧密
封面不得相对旋转，对所有螺丝联接件在装配时，均须涂抹润滑剂或油料调和的石墨，以免
咬死。阀门的使用：针形阀系线密封，仅需轻轻转动阀针，压紧密封面即能达到良好的密封
性能，禁止用力过大，以免损坏密封面。
5.超声波细胞破碎机安装好后，通入一定量的氮气保压30分钟，检查有无泄漏，如发现有泄
漏请用肥皂沫查找管路、管口泄漏点，找出后放掉气体拧紧，再次通入氮气保压试验，确保
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无泄漏后开始正常工作。
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6.当降温冷却时，可用水经冷却盘管进行内冷却，禁止速冷，以防过大的温差应力，造成冷
却盘管、釜体产生裂纹。工作时当釜内温度超过100℃时，磁力搅拌器与釜盖间的水套应通
冷却水，保证水温小于35℃，以免磁钢退磁。
7.安全装置：采用正拱型金属爆破片，材质为不锈钢，按国家标准GB567-89《拱型金属爆
破技术条件》制造，出厂时已试验好，不得随意调整。如果已爆破，需重新更换，更换期限
由使用单位根据本单位的实际情况确定，对于超过爆破片标定爆破压力而未爆破的应更换，
经常使用最好不超过爆破片的下限压力的80%，更换时应注意爆破片凸面向上。
8.反应完毕后，先进行冷却降温，再将釜内的气体通过管路泄放到室外，使釜内压力降至常
压，严禁带压拆卸，再将主螺栓、螺母对称地松开卸下，然后小心的取下釜盖(或升起釜盖)
置于支架上，卸盖过程中应特别注意保护密封面。
9.釜内的清冼：每次操作完毕用清洗液(使用清洗液应注意避免对主体材料产生腐蚀)清除釜
体及密封面的残留物，应经常清洗并保持干净，不允许用硬物质或表面粗糙的物品进行清洗。
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