CFSE说明书
CFSE染色
在使用前仔细阅读说明书-Cellstain-CFSE货号规格C375 1 mg荧光染料CFSE(CFDA-SE)，是一种可对活细胞进行荧光标记的新型染料，可以标记活体细胞。
其基本原理如下：CFSE是一种带有琥珀酰亚胺(NHS)的荧光染料，可以结合细胞内的蛋白质。
CFSE在结构上将酚羟基部位改造成AM体，所以自身不发荧光，荧光背景低，脂溶性高，能够轻易穿透细胞膜，在活细胞内与胞内蛋白共价结合，进入细胞内的CFSE 的AM部位能被细胞内酯酶水解发出绿色荧光。
CFSE的琥珀酰亚胺(NHS)和细胞内蛋白质的氨基部位结合，固定在细胞内，不会漏出细胞外。
CFSE进入细胞后定位于细胞膜、细胞质和细胞核，在细胞核的荧光最强。
在细胞分裂增殖过程中，CFSE的荧光强度会随着细胞的分裂而逐级递减，标记荧光可平均分配至两个子代细胞中，因此其荧光强度是亲代细胞的一半，根据这一特性，它可被用于检测细胞增殖，细胞周期的估算及细胞分裂等方面。
另外，CFSE标记的细胞用于体内观察可以长达数周之久，它常被用来做活体细胞检测实验和用荧光电镜观察细胞长期活动的实验。
有报道证实CFSE毒性小，不影响细胞的增殖能力。
此方法操作简单，且不用放射性同位素，不存在安全隐患。
可以更快速，更准确和更安全地得到想要的实验数据。
荧光波长：λex=496 nm, λem=516 nm储存条件运输条件-20℃室温所需设备) 移液器(20 μl, 1000 μl)培养箱(37 ℃CO2荧光显微镜(蓝色激发滤光片，绿色发射滤光片)流式细胞仪(488 nm光源，绿色发射滤光片)实验操作小鼠脾脏淋巴细胞分离与 CFSE 染色步骤实验材料：小鼠 1 只、组织剪 1 把、镊子 2 把、 75%酒精、小鼠固定板、1640 培养液（10%FBS， 1%双抗）、含 0.1%FBS 的 PBS 溶液（无菌）、PBS 溶液（无菌）、ACK 溶液（无菌）、移液管、枪头、组织研磨棒（无菌）、12 孔细胞培养板、15mL 离心管、300 目尼龙网（无菌），细胞计数板，手术台布。
CFSE原理和CFSE配制
细胞增殖检测：CFSE原理和CFSE配制分类：资料整理2011-08-08 10:08 阅读(27)评论(0)一、原理荧光染料CFSE，也可称为CFDA SE（5，6- carboxyfluorescein diacetate，succinimidyl ester）即羟基荧光素二醋酸盐琥珀酰亚胺脂，是一种可穿透细胞膜的荧光染料，具有与细胞特异性结合的琥珀酰亚胺脂基团和具有非酶促水解作用的羟基荧光素二醋酸盐基团，这使得CFSE成为一种良好的细胞标记物。
当CFSE 以含有两个乙酸基团和一个succinimidyl ester功能基团的形式存在时，不具有荧光性质，而具有细胞膜通透性，能够自由进入细胞；而当其扩散进入细胞内环境，内源的酯酶可将其乙酸基团水解，此种形式的CFSE分子具有很高的荧光活性，被激发能够产生绿色荧光，却不再具有膜通透性；同时，其含有的succinimidyl ester基团能与胞内的细胞骨架蛋白中的游离胺基反应，最终形成具有荧光的蛋白加合物。
因此，当细胞进行分裂增殖时，具有荧光的胞质蛋白被平均分配到第二代细胞中，这样与第一代细胞相比，其荧光强度便会减弱至一半；以此类推，分裂得到的第三代细胞的荧光强度便会比第二代细胞再次减弱。
这种现象可以在488nm的激发光下，采用流式细胞仪检测分析，通过检测到细胞荧光强度不断的降低，进一步分析得出细胞分裂增殖的情况。
二、CFSE配制用DMSO溶解成5 mmol/L的储存液，于- 20 ℃避光保存。
使用时，用无血清DMEM培养液稀释成5μmol/L的工作液备用。
CFSE试剂盒内有一小瓶CFSE（A 试剂）标明500ug ，另有一小瓶DMSO（B试剂），500ugCFSE溶解于180ulDMSO 即是5mMstock solution。
三、CFSE标记细胞制作细胞悬液，加入等体积CFSE工作液，于37℃孵育10 min，用40%体积的冷小牛血清立即终止标记10min。
Crossfire II 射程 glasses 产品说明书
PRODUCT MANUALThe Crossfire® II series of riflescopes offer the highestlevels of performance and reliability. With featuressuch as generous eye relief, rugged construction, andprecise, smooth controls, the Crossfire® II riflescopesare ready for any situation.Reticle FocusMagnificationAdjustment RingImages are for representation only.Product may vary from what is shown.Reticle Focal PlaneAll scope reticles can be termed either First FocalPlane (FFP) or Second Focal Plane (SFP) according to the reticle’s location within the scope. This model features an SFP reticle design.SFP reticles are located near the eyepiece, behind the image erecting and magnifying lenses. This style of reticle does not visually change in size when you change the magnification. The advantage of an SFP reticle is that it always maintains the same ideal visual appearance.Note: The Crossfire® II riflescope reticle subtensions used for bullet drop and wind drift compensation are correct at maximum magnification. The 6-24x50 model is correct at 18x magnification.Ocular FocusThe ocular focus is a one-time adjustment usedto focus the reticle for maximum sharpness. This adjustment is slightly different for every shooter. A clearly focused reticle is a critical component for accurate shooting.Ocular Focus – Reticle Focus AdjustmentYour Crossfire® II riflescope uses a Fast-Focus Eyepiece designed to quickly and easily adjust the focus on the riflescope’s reticle.To adjust the reticle focus:1. Set the magnification tothe highest setting. Ifequipped, set the parallaxto infinity.2. Back out the diopter untilthe reticle is slightly blurry.3. Turn the eyepiece in or out until the reticle imageis as crisp as possible.Note: Try to make this adjustment quickly, as the eye will try to compensate for an out-of-focus reticle.Once this adjustment is complete, it will not be necessary to refocus every time you use the riflescope. However, because your eyesight may change over time, you should recheck this adjustment periodically. Warning: Looking directly at the sun through a scope, or any optical instrument, can cause severe andpermanent damage to your eyesight.MagnificationThe magnification adjustment is used to change the riflescope’s magnification level, or “power,” adjusting from low to high magnification depending on the shooter’s preference.Magnification AdjustmentRotate the indicator bar to the desired magnification.TurretsThe Crossfire ® II riflescope incorporates adjustableelevation and windage turrets with audible clicks. Each audible click moves the bullet’s point of impact 1/4 of a minute of angle (MOA).1/4 MOA closely corresponds to 1/4 inch at 100 yards, 1/2 inch at 200 yards, and 3/4 inch at 300 yards. It takes four (4) clicks to move the bullet’s point of impact approximately one inch at 100 yards.To make turret adjustments:1. Remove the elevation and/or windage turret cap(s).2. Turn the turret in the direction you wish the bullet’spoint of impact to go: up or down, left or right.3. Replace the turret cap(s) when done.Tip: After sight-in, you can realign the zero marks on the elevation and windage turrets with the reference dots. See “Indexing the Turrets” section on page 15for instructions.Parallax is a phenomenon that results when the target image does not fall on the same optical plane as the reticle within the scope. This can cause an apparent movement of the reticle in relation to the target if theshooter’s eye is off-center.(Eye is not centered behind the scope.)•When the target image is not focused on the reticle plane and your eye is off-center behind the scope, parallax occurs. This is because the line of sight from the eye to the focused target image does not coincide with the reticle aiming point.(Eye is centered behind thescope.)• When the target image is not focused on the reticle plane and your eye is centered directly behind the scope, no parallax occurs. This is because the line of sight from the eye to the focused target image coincides with the reticle aiming point.(Eye is not centered behind the scope.)• When the target image is focused on the reticle plane, parallax cannot occur, even if your eye is not centered behind the scope. This is because the line of sight from the eye to the focused target image always coincides with the reticle aiming point no matter where the shooter’s eye is positioned.The Crossfire ® II riflescopes with fixed parallax are set for 100 yards. You will have a small amount of parallax inside and outside of 100 yards. Eliminate this by beingcentered perfectly behind the riflescope.Image Focus and Parallax CorrectionSome of the Crossfire ® II riflescope models use an image focus/parallax adjustment which providesmaximum image sharpness and eliminates parallax error. Lower power models do not use image focus/parallax adjustments and are pre-focused at 100 yards.Match shooting yardage to indicator dot. 100 yard setting shown.Adjustable Objective (AO models)Using image focus/parallax correction:1. Be sure the reticle is correctly focused (see ReticleFocus on page 5). 2. Rotate the adjustable objective until numbers matchthe distance you are shooting. Align yardage number to the indicator arrow on scope body.3. Check for proper setting by looking through thescope to verify image sharpness and, at the same time, look for reticle shift while moving your head back and forth.4. The setting is correct if there is no apparentmovement between the reticle and target while your head is moving back and forth. If there is apparent movement, adjust the focus knob slightly until the movement is eliminated.When properly set, the target image should be sharp and crisp.Note: The numbers on the adjustable objective are there for reference and may perfectly correspond with the actual yardage.Reticle Illumination AdjustmentSome Crossfire® II riflescope models offer anilluminated reticle controlled by an adjustment knob on the eyepiece. Adjust the illumination intensity by rotating the knob either clockwise or counter-clockwise. Battery Replacement1. Unscrew the outer cap with a coin.2. Remove the battery.3. Replace with a new CR 2032 battery.4. Install battery with plus (+) side up.5. Reinstall the outer cap and tighten securely.Battery cover on top ofadjustment knob.To get the best performance from your Crossfire® II riflescope, proper mounting is essential. Although not difficult, the correct steps must be followed. If you are unsure of your abilities, use the services of aqualified gunsmith.Rings And BasesMount an appropriate base and matching rings to your rifle according to the manufacturer’s instructions. Your new Crossfire® II riflescope requires either 1-inch or 30mm rings. Refer to the dimensions listed on the box or go to and look up your model and configuration to confirm the main tube size for your riflescope.Use the lowest ring height that will provide complete clearance of scope and rifle, avoiding any contact with barrel, receiver, bolt handle, or any other part of the rifle. A low mounting will help assure proper cheek weld, aid in establishing a solid shooting position, and promote fast target acquisition.Eye Relief And Reticle AlignmentInstall the bottom ring halves on the mounting base. Place the scope on the bottom ring halves and loosely install the upper ring halves. Before tightening the scope ring screws, adjust for comfortable eye relief:1. Set the scope to maximum magnification.2. Slide the scope as far forward in the ringsas possible.3. Look through the scope while in your normalshooting position and slowly slide the scope towards your eye. Stop sliding the scope when you see thefull field of view.Note: Keep the rings centered on the riflescope tube asmuch as possible. You may have to adjust where the rings are mounted on the rail to get both the correct mount andproper eye relief.4. Without disturbing the front-back placement, rotatethe scope until there is an exact match betweenthe reticle’s vertical crosshair and the rifle’s vertical axis. Use a reticle leveling tool, a weight hung ona rope, flat feeler gauges, or a bubble level to helpwith this procedure.Note: After aligning the reticle, tighten and torque the ring screws. Vortex Optics recommends a torque setting of 15-18 in/lbs on the ring screws. DO NOT use a thread lockingcompound on the threads. Thread locking agents lubricate the threads, which can increase the applied torque.Use of bubble levels to square the riflescope(and reticle) to the base.Bore SightingInitial bore sighting of the riflescope will save time and money at the range. You can do this in a number of ways. Use a mechanical or laser bore sighter according to the manufacturer’s instructions. On some rifles, you can bore sight by removing the bolt and sighting throughthe barrel.To Visually Bore Sight A Rifle:1. Place the rifle securely on a rest and removethe bolt.2. Sight through the bore at a target approximately100 yards away.3. Move the rifle and rest until the target is visuallycentered inside the barrel.4. With the target centered in the bore, make windageand elevation adjustments until the reticle crosshair is also centered over the target.Final Range Sight-InAfter the riflescope has been bore sighted, you should do your final sight-in at the range, using the exact ammunition you expect to use while shooting. Sight-in and zero the riflescope at the preferred distance. 100 yards is the most common zero distance, although you may prefer a 200-yard zero for long-range applications.• Following all safe shooting practices, fire a three-shot group as precisely as possible.• Next, adjust the reticle to match the approximate center of the shot group (see section on Windage and Elevation Adjustment on page 5).Note: If the rifle is securely mounted and cannot be moved, simply look through the scope and adjust the reticle until it is centered on the group.• Carefully fire another three-shot group and see if the bullet group is centered on the bullseye.Repeat this procedure as many times as necessary to achieve a perfect zero.Indexing the TurretsThe Crossfire® II riflescopes feature windage/elevation dials that allow you to reindex the zero indicator after sight-in without disturbing your settings. Though not required, this process will allow you to quickly return to your original zero if you dial temporary corrections in the field.Index the turret as follows:1. Remove the turret caps.2. While firmly holding thedial, loosen and removecenter screw.3. Lift the turret dial offthe scope. Align thezero on the turret dialwith the index line.4. Reinstall the turret dialand tighten the centerscrew while holdingfirmly to the turret dial.CleaningThe Crossfire® II riflescope requires very little routine maintenance other than periodically cleaning the exterior lenses. Clean the scope’s exterior by wiping with a soft, dry cloth. When cleaning the lenses, be sure to use products specifically designed for use on coated optical lenses.• Be sure to blow away any dust or grit on the lenses prior to wiping the surfaces.• Using your breath, or a very small amount of water or pure alcohol, can help remove stubborn things like dried water spots.• Check out our cleaning kits at LubricationAll components of the Crossfire® II are permanently lubricated, so no additional lubricant should be applied. If possible, avoid exposing your riflescopeto direct sunlight or any very hot location for long periods of time.Note: Other than removing the turret caps and battery cap, do not attempt to disassemble any scope components. Disassembling the scope may void the warranty.StorageIf possible, avoid exposing your riflescope to direct sunlight or any very hot location for long periods of time.Sighting-In ProblemsMany times, problems thought to be with the scope are actually mount problems. Be sure you’re using the correct base and rings in the correct orientation, and the base screws and rings are tight. Insufficient windage or elevation adjustment range may indicate problems with rings, base, base alignment, or barrel/ receiver alignment.Check For Correct Base And Ring Alignment • Roughly center the reticle by adjusting both windage and elevation turrets to the midpoint of their travel ranges.• Attach bore sighter or remove bolt and visually boresight rifle.• Look through the scope. If the reticle appearsway off-center on the bore sighter image, or when compared to the visually centered target when looking through rifle’s bore, there may be a problem with the bases or rings being used. Confirm you are using the correct base and rings in the proper orientation.Tips For Solving Bullet Grouping Problems• Maintain a good shooting technique and use a solid rest.• Check that all screws on rifle’s action are properly tightened.• Be sure rifle barrel and action are clean and free of excessive oil or copper fouling.• Check that rings are correctly torqued per the manufacturer’s instructions.• Some rifles and ammunition don’t work well together—try different ammunition and see if accuracy improves.VIP WARRANTYOUR UNCONDITIONAL PROMISE TO YOU.Learn more at ************************•800-426-0048NOTE: The VIP Warranty does not cover loss, theft, deliberate damage, or cosmetic damage not affectingproduct performance.For additional and latest manuals, visit We promise to repair or replace the product. Absolutely free.Unlimited. Unconditional. Lifetime Warranty.M-00214-1© 2020 Vortex Optics® Registered Trademark and TM Trademark of Vortex Optics。
碳硫分析仪使用说明书
前言欢迎使用我公司的仪器，它将给您的分析工作带来许多方便，为您创造良好的经济效益。
我公司会提供优质可靠的服务。
您在使用仪器前请耐心阅读本使用说明书，尤其要注意安全要求！如果仪器分析部分是气容仪，第二章第二节可不读；如果仪器分析部分是滴定仪，第二章第一节可不读。
JN8608-XX型电弧燃烧碳硫分析仪是一种系列产品。
它使用燃烧法分析碳、硫成分的百分含量，最适合于对钢铁样品的分析，在加入一定添加剂情况下，也可对其它样品（如赤铁矿、硅矿、锰矿、炉渣、焦炭等）碳硫成分进行定量分析。
该仪器由燃烧部分和分析部分组成。
由于两部分有一些不同的特点，又区分不同的一些序列号。
本说明书适合于JN8608-1B、JN8608-1C、JN8608-2B、JN8608-2C、JN8608-2D、JN8608-2E、JN8608-2F这几个型号的仪器，对上述型号的仪器通称为JN8608-XX电弧燃烧碳硫分析仪。
仪器燃烧部分叫做电弧炉，其分析部分有气体容量仪（简称气容仪），滴定仪之区分，或其它检测碳硫的设备。
其型号代码如下：JN8608-X X 金牛高速分析仪器有限公司电弧燃烧碳硫分析仪分析方法特征号：1、代表非水分析法2、代表气体容量法分析部分特征号：2B气容仪标尺直读（简称2B型气容仪）2C气容仪数字显示（简称2C型气容仪）2D气容仪数字显示，自动打印测试结果（简称2D型气容仪）2E气容仪数字显示，硫自动滴定（简称2E型气容仪）2F气容仪数字显示，工作过程全自动控制，自动打印测试结果（简称2F型气容仪）1B滴定仪气压加液（简称1B型滴定仪）1C滴定仪手压加液（简称1C型滴定仪）本公司产品常有所改进，仪器的某些部分可能与说明书不一致，敬请用户原谅。
目录第一章燃烧部分——电弧炉 (1)第一节概述 (1)第二节电弧燃烧炉技术参数 (1)第三节电弧燃烧炉操作方法 (1)第四节操作须知 (3)第五节常见主要故障及其排除 (5)第二章分析部分 (6)第一部分、气容仪 (6)第一节概述 (6)第二节主要技术性能 (7)第三节基本工作原理 (7)第四节操作方法 (10)第五节计量定标 (12)第六节仪器漏气检测方法 (12)第七节仪器调试 (12)第八节常见故障的排除方法 (13)第二部分、滴定仪 (15)第一节概述 (15)第二节测量范围 (15)第三节结构及使用说明 (15)第四节溶液配制 (15)第五节操作方法 (17)第六节使用注意事项 (18)第一章燃烧部分——电弧炉一、概述JN8608电弧燃烧碳硫分析仪燃烧部分称为电弧燃烧炉（以下简称电弧炉)，在用燃烧法分析物质成分时，本电弧炉用于燃烧样品，可将其燃气导入各种分析设备，定量分析样品中碳、硫含量。
CFDA SE (细胞增殖示踪荧光探针) 说明书
CFDA SE (细胞增殖示踪荧光探针) 产品编号产品名称包装C1031 CFDA SE (细胞增殖示踪荧光探针) 5mg产品简介：CFDA SE 的全称为Carboxyfluorescein diacetate, succinimidyl ester ，是一种近年来被广泛应用的细胞增殖检测用荧光探针，也可以用于细胞的荧光示踪。
基于CFDA SE 荧光标记的细胞增殖检测和[3H]-thymidine 掺入、BrdU 标记获得的检测结果完全一致，但同时可以提供更多的细胞增殖信息。
使用CFDA SE 检测可以提供整个细胞群中有多少比例的细胞分裂了1次、2次或更多次数，同时如果和其它荧光探针联用，可以获取不同分裂次数细胞的其它相关信息。
CFDA-SE 的分子式为C 29H 19NO 11，分子量为557.47，CAS number 为150347-59-4。
CFDA SE 可以通透细胞膜，进入细胞后可以被细胞内的酯酶(esterase)催化分解成CFSE ，CFSE 可以偶发性地(spontaneously)并不可逆地和细胞内蛋白的Lysine 残基或其它氨基发生结合反应，并标记这些蛋白。
在加入荧光探针CFDA SE 后大约24小时，即可充分标记细胞。
被CFDA SE 标记的非分裂细胞的荧光非常稳定，稳定标记的时间可达数个月。
CFDA SE 标记细胞的荧光非常均一，比以前使用的其它细胞示踪荧光探针例如PKH26的荧光更加均一，并且分裂后的子代细胞的荧光分配也更均匀。
由于CFDA SE 标记细胞的荧光非常均匀和稳定，每分裂一次子代细胞的荧光会减弱一半，这样通过流式细胞仪检测就可以检测出没有分裂的细胞，分裂一次的细胞(1/2的荧光强度)，分离两次的细胞(1/4的荧光强度)，分裂三次的细胞(1/8的荧光强度)以及类似的其它分裂次数的细胞。
采用CFDA SE 通过流式细胞仪检测获得的检测结果参考右图。
每一个峰代表一种分裂次数的细胞，从右至左的峰通常依次为分裂0次、1次、2次、3次等次数的细胞。
芬尼斯蒸汽手持蒸汽器说明书
Handheld garment steamer1000W, up to 20 g/minVertical Steaming70ml Detachable water tankGC350/46Compact solution for easy de-wrinklingIdeal companion to your iron, no board requiredThis handheld garment steamer is perfect for delicate fabrics, difficult-to-iron areas and torefresh any clothing or upholstery - no burns guaranteed. Lightweight and compact designmakes it easy to use anytime anywhere. Just steam and go!Quick de-wrinkling•Automatic continuous steam for easy de-wrinkling•Brush accessory for thicker fabricsEasy to use and safe•Safe on all ironable fabrics, no burns guaranteed•No ironing board needed•Compact design for convenient use, storage and travel•Detachable water tank for easier filling•2.5m cord for longer reachRefreshes clothes without washing or dry cleaning•Continuous steam removes odors and kills 99.9%* of bacteriaIssue date 2022-03-30Version: 1.1.2EAN: 08 71010 38339 94© 2022 Koninklijke Philips N.V.All Rights reserved.Specifications are subject to change without notice. Trademarks are the property of Koninklijke Philips N.V. or their respective SpecificationsHandheld garment steamer1000W, up to 20 g/min Vertical Steaming, 70ml Detachable water tankHighlightsContinuous steamAn electric pump automatically provides continuous steam for easy and quick de-wrinkling.Brush accessoryThe brush attachement opens the fabric fibers and enables better steam pemetration. It is especially good for thicker garments like jackets and coats. It can also help remove dirts and pills.Safe on all ironable fabricsThe steamer is safe to use on all ironable fabrics and garments. The steam plate can be safely pressed against any clothing with no risk of burning – a great solution for delicate fabrics, like silk.No ironing board neededUsing a clothes steamer on hanging garments eliminates the need for an ironing board, so de-wrinkling is easy and hassle-free.Ergonomic designThe handheld garment steamer is ergonomically designed to be light, compact and comfortable to use. Just press the trigger and watch wrinkles and creases disappear.Detachable water tankThe water tank detaches for easy filling under the tap.2.5m cordfor longer reach99.9% bacteria-free*Hot steam refreshes your clothes and kills up to 99.9% of bacteria*. Less frequent washing and dry cleaning saves time and money, and helps clothes last longer.Easy to use•Safe on all ironable fabrics: Even delicates like silk •Water tank capacity: 70 ml •Detachable water tank •Refill any time during use •Power cord length:2.5 m •Ready to use: Light indicatorAccessories included•Brush•Glove for extra protectionFast crease removal•Power: 1000 W•Continuous steam: Up to 20 g/min •Voltage: 220-240 V•Ready to use: <1 minute(s)Size and weight•Packaging dimensions (WxHxL): 38 x 12.8 x 15 cmGreen efficiency•Product packaging: 100% recycable •User manual: 100% recycled paperConsumer Trade Item•Height:15•Width: 38•Length: 12.8•Net Weight: .793•Gross Weight: 1.01•GTIN: 08710103833994•Country of origin: CN•Harmonized system code: 851679Outer Carton•Height:32•Width: 27.1•Length: 39.5•Gross Weight: 4.685•GTIN: 18710103833991Guarantee•2 year worldwide guarantee*Tested by external body for bacteria types Escherichia coli 8099, Staphylicoccus aureus ATCC 6538, Canidia albicans ATCC 10231 with 1 minute steaming time.。
ROS活性氧诱导剂
彻底清除残留清洁剂。 6. 实验后完成后所有样品及接触过的器皿应按照规定程序处理。
组织活性氧检测试剂盒（O13）
组织
Байду номын сангаас
冰冻切片活性氧检测试剂盒（O11） 冰冻切片
组织活性氧检测试剂盒（O11）
组织
冰冻切片活性氧检测试剂盒（O13） 冰冻切片
组织活性氧检测试剂盒（O12）
组织
冰冻切片活性氧检测试剂盒（O12） 冰冻切片
荧光颜色 绿色 红色 绿色 绿色 绿色 绿色 绿色 绿色 红色 绿色 绿色
产品说明书
ROS 活性氧诱导剂 / ROS 诱导剂
货号：BB-47058
V 8.9
试剂盒储存条件：
-20℃ 密封避光保存。
【注】： 开盖后组份按要求条件保存。 拆封后请尽快使用完！
试剂盒组成：
产品组成
BB-47058-1 BB-47058-2
规格
1 ml
5 ml
ROS 活性氧诱导剂 / ROS 诱导剂（1000X）
产品号 BB-4104 BB-4105 BB-4106 BB-4107 BB-4108 BB-4112 BB-4181 BB-4123 BB-4137 BB-4132 BB-4221 BB-44111 BB-44120 BB-44138 BB-470512 BB-4731 BB-48120 BB-48131 BB-48121
1 ml
1 ml*5
使用说明书
【求助】荧光染料CFSE的配置与使用
【求助】荧光染料CFSE的配置与使用这是我根据同仁的说明书和网上查到的资料总结会总后的内容，你可以参考一下Cellstain- CFSE一羧基荧光素二醋酸盐琥珀酰亚胺酯羟基荧光素二醋酸盐琥珀酰亚胺脂(CFSE) 是一种可穿透细胞膜的荧光染料,具有与细胞特异性结合的琥珀酰亚胺脂基团和具有非酶促水解作用的羟基荧光素二醋酸盐基团,使CFSE 成为一种良好的细胞标记物[1 ] 。
CFSE 进入细胞后可以不可逆地与细胞内的氨基结合偶联到细胞蛋白质上。
当细胞分裂时,CFSE 标记荧光可平均分配至两个子代细胞中,因此其荧光强度是亲代细胞的一半。
这样,在一个增殖的细胞群中,各连续代细胞的荧光强度呈对半递减,利用流式细胞仪在488nm 激发光和荧光1(FL1) 检测通道可对其进行分析Product Code: C375-10 Unit: 1 mg ，白或淡黄色粉末保存：－20℃ 运输：室温CFSE的配制和细胞染色程序DMSO是有机溶剂，对于蛋白质会有损伤，所以在配置的时候DMSO溶液越少越好；CFSE遇水容易分解，所以在配置时候要用无水DMSO，并且配置后尽快试验。
1、离心：3000rpm ?10min；2、取DMSO 180ul，稀释CFSE，超音波溶解，得到10mM CFSE保存液；将其按20ul体积分装于预遮光的无菌冻存管中，共分9管，标记后其中8管保存于－20℃。
另一管按10ul装于管中保存。
保存2个月。
3、取上余之10ul保存液，用PBS稀释至50ul，得到5mM的CFSE稀释液，保存于－20℃待用。
CFSE 标记细胞{KJ, 2002 #1}:标记浓度和条件：细胞通常用终浓度0.5－5μM的CFSE标记。
孵育时间5－10分钟，通常在10％FCS的PBS中染色，标记后细胞用完全培养介质洗涤，高蛋白灭活未反应的CFSE。
1、在有5％FCS的PBS重悬细胞，细胞浓度一般为1?106 （体外实验）?107/ml（转染实验）。
CFSE使用方法
CellTrace ™ Cell Proliferation KitsCatalog nos. C34554, C34557, C34564IntroductionThe CellTrace ™ Cell Proliferation Kits provide versatile and well-retained cell tracing reagents in a convenient and easy-to-use form. Each kit contains a CellTrace ™ reagent (CellTrace ™ CSFE, CellTrace ™ Violet, or CellTrace ™ Far Red) in single-use vials to permit small scale experiments without preparing excess quantities of stock solution. TheCellTrace ™ reagents easily diffuse into cells and bind covalently to intracellular amines, resulting in stable, well-retained fluorescent staining that can be fixed with aldehyde fixatives. Excess unconjugated reagent passively diffuses to the extracellular medium, where it can be quenched with complete media and washed away.Spectral Characteristics CellTrace ™ CSFE (Cat. no. C34554)The approximate excitation and emission peaks of this product after hydrolysis are 492 nm and 517 nm, respectively. Cells labeled with CellTrace ™ CSFE reagent can be visualized by fluorescence microscopy using standard fluorescein filter sets or analyzed by flow cytometry in an instrument equipped with a 488-nm excitation source.CellTrace ™ Violet (Cat. no. C34557)The approximate excitation and emission peaks of this product after hydrolysis are 405 nm and 450 nm, respectively. Cells labeled with CellTrace ™ Violet reagent can be visualized by fluorescence microscopy using standard DAPI filter sets or analyzed by flow cytometry in an instrument equipped with a 405-nm excitation source.CellTrace ™ Far Red (Cat. no. C34564)The approximate excitation and emission peaks of this product after hydrolysis are 630 nm and 661 nm, respectively. Cells labeled with CellTrace ™ Far Red reagent can be visualized by fluorescence microscopy using standard Cy ®5 filter sets or analyzed by flow cytometry in an instrument equipped with a 633/635-nm excitation source.Table 1Contents and storageBefore StartingMaterials Required but NotProvided• Cells of interest as a single-cell suspension• Phosphate-buffered saline (PBS) or similar protein-free buffer • Culture media containing protein such as FBS or BSA • Flow cytometerCautionNo data are available addressing the mutagenicity or toxicity of CellTrace ™ reagents (Component A). Handle the DMSO dye solution with caution because DMSO is known to facilitate the entry of organic molecules into tissues. Always wear suitable protective clothing, gloves, and eye/face protection when handling this reagent. Dispose of the reagents in compliance with all pertaining local regulations.Storage and HandlingUpon receipt, store the kit components desiccated at ≤–20°C until required for use. When stored properly DMSO and dry CellTrace ™ reagents are stable for at least one year. Allow the product to warm to room temperature before opening vials. Stocksolutions of CellTrace ™ reagents should be used only on the day of preparation. Avoid repeated freezing and thawing of kit contents.Figure 1 Generational tracing using CellTrace ™ Reagents. (A) Cell proliferation was followed for 8 generations using the CellTrace ™ Violet reagent. Human peripheral blood mononuclear cells were harvested and stained with CellTrace ™ Violet reagent prior to stimulation with mouse anti-human CD3 (Cat. no. MHCD0300) and Interleukin-2 (Cat. no. PHC0027) for 7 days. The discrete peaks in this histogram represent successive generations of live, CD4 positive cells. Analysis was completed using an Attune ® Acoustic Focusing Cytometer with 405-nm excitation and a 450/40 bandpass emission filter . The unstimulated parent generation is indicated in red. (B) and (C) Cell Proliferation was followed for 7 generations using the CellTrace ™ CFSE and CellTrace ™ Far Red reagents, respectively. Human T lymphocytes were harvested and stained with CellTrace ™ CFSE or CellTrace ™ Far Red reagent prior to stimulation with anti-human CD3 (Cat. no. MHCD0300) for 5 days. The discrete peaks in these histograms represent successive generations of live, SYTOX ® Green (Cat. no. S34860) negative cells. The unstimulated parent generation is indicated in blue (CellTrace ™ CFSE) or purple (CellTrace ™ Far Red). Analysis was completed using an Attune ® Acoustic Focusing Cytometer with 488 nm excitation and a 530/30 nm bandpass emission filter for CellTrace ™ CFSE. Analysis was completed for CellTrace ™ Far Red using an Attune ® Acoustic Focusing Cytometer with 638 nm excitation and a 660/20 nm bandpass emission filter.CellTrace ™ CFSE Fluorescence P e r c e n t o f M ax103104105106801004020060CellTrace ™ Violet FluorescenceC e l l C o u n t10310410510620001000102400003000CellTrace ™ Far Red FluorescenceP e r c e n t o f M a x103104105106801004020060ABCExperimental ProtocolsGeneral Guidelines• The following methods have been optimized for monitoring cell proliferation of human T and B lymphocytes.• In other cell types and applications, we recommend titration of the CellTrace ™reagents to determine the optimal staining concentration. CellTrace ™ stock solutions may be diluted in DMSO for this purpose.• Recommended concentration for CellTrace ™ staining is 1–10 μM. A dye concentration of 5–10 μM is recommended for tracking five or more generations, while 1–2 μM may be sufficient for tracking less than five generations.• Start with a single cell suspension in PBS for uniform cell labeling.• Uniform cell labeling can also be improved by gently agitating cells during staining.• A low flow rate should be used for analysis on hydrodynamic focusing cytometers to ensure separation of distinct generational peaks. All collection rates will provide equivalent results when analyzing on an Attune ® Acoustic Focusing Cytometer.• To ensure appropriate instrument setup, include a sample of unstimulated control cells in proliferation experiments using CellTrace ™ reagents.Table 2 Preparation of CellTrace ™ stock solutionsLabeling Cells in SuspensionThe following protocol has been optimized for cell concentrations of up to 106 cells/mL. Dye concentration may need to be increased for samples with >106 cells/mL.1.1 Prepare CellTrace ™ stock solution immediately prior to use by adding the appropriatevolume of DMSO (Component B) to one vial of CellTrace ™ reagent (Component A) and mixing well (see Table 2, above). 1.2 Add 1 µL of CellTrace ™ stock solution in DMSO (prepared in Step 1.1) to each mL of cellsuspension in PBS for a final working solution (see Table 2, above, for concentration). 1.3 Incubate the cells for 20 minutes at room temperature or 37°C, protected from light.1.4 Add five times the original staining volume of culture medium (containing at least1% protein) to the cells and incubate for 5 minutes. This step removes any free dye remaining in the solution.1.5 Pellet the cells by centrifugation and resuspend them in fresh pre-warmed completeculture medium.1.6 Incubate the cells for at least 10 minutes before analysis to allow the CellTrace ™ reagentto undergo acetate hydrolysis. 1.7 Proceed with cell stimulation, incubation, or analysis.Alternate Method for LabelingCells in Suspension The following protocol has been optimized for cell concentrations up to 106 cells/mL.Dye concentration may need to be increased for samples with >106 cells/mL.2.1 Prepare CellTrace™ stock solution immediately prior to use by adding the appropriatevolume of DMSO (Component B) to one vial of CellTrace™ reagent (Component A) andmixing well (see Table 2, page 3).2.2 Pellet the cells by centrifugation and remove the supernatant.2.3 Dilute the CellTrace™ DMSO stock solution in pre-warmed (37°C) phosphate-bufferedsaline (PBS) or other protein-free buffer to the desired working concentration (1–10 µM).2.4 Gently resuspend the cells in the PBS dye solution (prepared in Step 2.1).2.5 Incubate the cells for 20 minutes at room temperature or 37°C, protected from light.2.6 Add five times the original staining volume of culture medium (containing at least1% protein) to the cells and incubate for 5 minutes. This step removes any free dyeremaining in the solution.2.7 Pellet the cells by centrifugation and resuspend them in fresh, pre-warmed completeculture medium.Incubate the cells for at least 10 minutes before analysis to allow the CellTrace™ reagent2.8to undergo acetate hydrolysis.Proceed with cell stimulation, incubation, or analysis.2.9Alternate Method for LabelingAdherent Cells3.1 Prepare CellTrace™ stock solution immediately prior to use by adding the appropriatevolume of DMSO (Component B) to one vial of CellTrace™ reagent (Component A) andmixing well (see Table 2, page 3).3.2 Grow the cells to the desired density on coverslips or flasks filled with the appropriateculture medium.3.3 Dilute the CellTrace™ DMSO stock solution in pre-warmed (37°C) phosphate-bufferedsaline (PBS) or other protein-free buffer to the desired working concentration (1–10 µM).This is the loading solution.3.4 Remove the culture medium from the cells and replace it with the loading solution(prepared in Step 3.3).3.5 Incubate the cells for 20 minutes at 37°C.3.6 Remove the loading solution, wash the cells twice with culture medium containing atleast 1% protein, and replace with fresh, pre-warmed complete culture medium.Incubate the cells for at least 10 minutes before analysis to allow the CellTrace™ reagent3.7to undergo acetate hydrolysis.Proceed with cell stimulation, incubation, or analysis.3.8Optional: Fixation andPermeabilizationLabel the cells with CellTrace™ reagent according to one of the protocols listed above.4.14.2 Before fixation, wash and resuspend the cells in PBS or other protein-free buffer.4.3 Fix the cells for 15–20 minutes at room temperature using an aldehyde-based fixativesuch as paraformaldehyde, protected from light.4.4 Wash the cells with PBS.4.5 If desired, permeabilize the cells using any appropriate protocol. CellTrace™ reagentscovalently bind to cells and will not wash out after permeabilization.4.6 Following permeabilization, wash the cells with PBS.4.7 Resuspend the cells in PBS prior to analysis.Combining CellTrace™Reagents with otherFluorescent Markers5.1 Label the cells with CellTrace™ reagent according to one of the protocols listed above.5.2 Resuspend the cells in a buffer appropriate for the subsequent staining applications (seeProduct List, below).5.3 Apply stains for immunophenotyping, DNA content, apoptosis, or other markers asrecommended for each stain.References1. J Cell Biol 101, 610 (1985);2. J Cell Biol 103, 2649 (1986);3. J Immunol Methods 171, 131 (1994);4. J Exp Med 184, 277 (1996);5. J Immunol Methods 133, 87 (1990);6. Transplant Proc 24, 2820 (1992);7. Current Protocols in Cytometry, J. P. Robinson, Ed., (1998) pp 9.11.1-9.11.9.;8. Cytometry Part A, 79A: 95–101 (2011); 9. Current Protocols in Immunology, R. Coico, Ed., (2008) pp 7.10.1-7.10.24.10 June 2014Purchaser NotificationThese high-quality reagents and materials must be used by, or directl y under the super v ision of, a tech n ically qualified individual experienced in handling potentially hazardous chemicals. Read the Safety Data Sheet provided for each product; other regulatory considerations may apply.Obtaining SupportFor the latest services and support information for all locations, go to .At the website, you can:• Access worldwide telephone and fax numbers to contact Technical Support and Sales facilities • Search thr ough frequently asked questions (FAQs)• Submit a question directly to Technical Support (techsupport@ )• Search for user documents, SDSs, vector maps and sequences, application notes, formulations, handbooks, certificates of analysis, citations, and other product support documents• Obtain information about customer training • Download software updates and patchesSDSSafety Data Sheets (SDSs) are available at /sds .Certificate of AnalysisThe Certificate of Analysis provides detailed quality control and product qualification information for each product. Certificates of Analysis are available on our website. Go to /support and search for the Certificate of Analysis by product lot number, which is printed on the product packaging (tube, pouch, or box).Limited Product WarrantyLife Technologies Corporation and/or its affiliate(s) warrant their products as set forth in the Life Technologies’ General Terms and Conditions of Sale found on Life Technologies’ website at /termsandconditions . If you have any questions, please contact Life Technologies at /support .DisclaimerLIFE TECHNOLOGIES CORPORATION AND/OR ITS AFFILIATE(S) DISCLAIM ALL WARRANTIES WITH RESPECT TO THIS DOCUMENT, EXPRESSED OR IMPLIED, INCLUDING BUT NOT LIMITED TO THOSE OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE, OR NON-INFRINGEMENT. TO THE EXTENT ALLOWED BY LAW, IN NO EVENT SHALL LIFE TECHNOLOGIES AND/OR ITS AFFILIATE(S) BE LIABLE, WHETHER IN CONTRACT, TORT, WARRANTY, OR UNDER ANY STATUTE OR ON ANY OTHER BASIS FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE OR CONSEQUENTIAL DA MAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT, INCLUDING BUT NOT LIMITED TO THE USE THEREOF.Important Licensing InformationThese products may be covered by one or more Limited Use Label Licenses. By use of these products, you accept the terms and conditions of all applicable Limited Use Label Licenses.All trademarks are the property of Thermo Fisher Scientific and its subsidiaries, unless otherwise specified. Cy is a registered trademark of GE Healthcare UK Limited. ©2014 Thermo Fisher Scientific Inc. All rights reserved.Product List Current prices may be obtained from our website or from our Customer Service Department.Cat. no. Product Name Unit SizeC34554 CellTrace ™CSFE Cell Proliferation Kit *for flow cytometry*...................................................... 1 kitC34557 CellTrace ™Violet Cell Proliferation Kit *for flow cytometry*......................................................1 kit C34564 CellTrace ™Far Red Cell Proliferation Kit *for flow cytometry*.................................................... 1 kit Related ProductsMHCD0300 Purified Mouse anti-human CD3 ............................................................................ 500 μL PHC0026 Recombinant Human Interleukin-2.......................................................................... 10 μgS10274 SYTOX ® AADvanced ™Dead Cell Stain Kit *500 tests* ........................................................ 5 × 0.5 mL S10349 SYTOX ® AADvanced ™ Dead Cell Stain Kit *100 tests* ........................................................... 0.2 mLS34859 SYTOX ® Red Dead Cell Stain *for flow cytometry* *1000 tests*...................................................1 mL S34860 SYTOX ®Green Dead Cell Stain *for flow cytometry* *1000 tests* ................................................. 1 mL111.31D DynaBeads ®CD3/CD28 T Cell Expander...................................................................... 2 mL08-0022SA OpTmizer ™T-Cell Expansion SFM........................................................................... 500 mL 10439-016 Fetal Bovine Serum, ES Cell-Qualified ....................................................................... 100 mL 14190-136 Dulbecco’s Phosphate-Buffered Saline (D-PBS) (1X), liquid...................................................... 1000 mL。
CFSE原理和CFSE配制
CFSE原理和CFSE配制CFSE（Crystal Field Splitting Energy）是晶场分裂能的缩写，也被称为晶场裂化能。
它是指发生在过渡金属离子周围配位体形成晶场的时候，能级发生分裂的能量差。
过渡金属离子在担任催化剂或作为配体参与反应时，其电子结构的变化对反应的速率和选择性有重要影响。
CFSE可以解释过渡金属配合物的颜色、磁性、还原性等性质。
CFSE原理的基础来自晶场理论，该理论认为配位体中的电子对过渡金属离子周围形成六个配位点，组成一个八面体。
当八个配位体完全靠近过渡金属离子时，它们与金属离子之间会产生相互作用，形成一个晶场。
这个晶场将过渡金属离子的d轨道分成两个部分，等效于低能级或t2g轨道和高能级或eg轨道。
在这个晶场的作用下，原先能量均匀分布的五个d轨道被分成了两组。
低能级组（t2g轨道）中有三个轨道，高能级组（eg轨道）中有两个轨道。
这种能级分裂称为晶场分裂或晶场裂化。
CFSE配制是指通过改变配位体的种类、配位数或者配位体与金属之间的配位方式，来调控晶场分裂能。
常用于CFSE配制的方法包括合成新的配合物、改变反应条件以及使用不同的反应溶剂。
通过调控晶场分裂能，可以改变过渡金属离子的电子结构，进而影响配合物的性质。
例如，如果把晶场分裂能增大，即使配位体提供了足够的电子给金属离子，也无法填满所有的t2g轨道，这就使得高能级的轨道变得更加稳定。
结果是，配位体在吸收光子时，只能吸收较低能量的光，这就使得配合物呈现出较深的颜色。
另一方面，通过改变配位体和过渡金属离子的配位方式，也可以改变CFSE分裂模式。
例如，一种是四面体配位模式，即配位体以四面体的形式围绕过渡金属离子。
在这种情况下，晶场分裂使得t2g轨道和eg轨道之间的能隙更小，即晶场分裂能降低。
这种分裂模式可以导致配合物呈现出无色或者较浅颜色。
总之，CFSE原理和CFSE配制可以帮助我们理解和控制过渡金属离子的电子结构，进而影响过渡金属配合物的性质。
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CellTrace™ CFSE 细胞增殖试剂盒
贮存条件：
• ≤–20ºC 避免反复冻融，开盖前放置至室温。
• 干燥
• 避光
按要求保存，DMSO和固体CFSE6个月稳定，试剂溶液尽快使用。
分子量：557.47
Ex/Em:492/517 nm
检测：荧光显微镜FITC滤片检测或流式488nm激发光检测。
CFSE (carboxyfluorescein diacetate, succinimidyl ester 羧基荧光素双乙酸盐，琥珀酰亚胺酯) 被动扩散进入细胞，其本身是无色无荧光的，被细胞内的酯酶分解后产生高强度的荧光，该荧光产物与胞内的氨基结合长时间留存于细胞内并可被乙醛固定。
未结合的试剂与副产物扩散至胞外基质中，被洗去。
传代或胚胎分裂分析，CFSE与赖氨酸侧链或其他氨基基团反应，共价偶联至细胞内和细胞表面的蛋白，细胞分裂时被平均分配至子代细胞，荧光强度降到母代细胞的一半。
适合用于胞内示踪，在细胞分裂或融合后分配至子代细胞中，并不会被转移至邻近的细胞。
在小鼠淋巴细胞迁移实验中，CFSE注射进入小鼠体内后，标记的淋巴细胞的检测可长达8周。
肝内注射荧光显微镜定位检测可长达20天。
试剂盒成分
• CellTrace™ CFSE (Component A), 10瓶，每瓶50 μg
• DMSO (Component B), 0.5 mL
使用浓度：
一般来说，长时间染色（超过三天）或快速分裂为5～10uM，活力分析等短时间分析为0.5～5uM。
，荧光显微镜为25uM。
优化并使用尽可能低的浓度以保持细胞的正常生理状态。
不要使用含氨基的缓冲液或赖氨酸涂层的玻片，防止和CFSE发生反应。
试剂准备：
用前制备CFSE的5mM贮存液，一瓶A溶于18ul DMSO（B）中。
流式分析：
已用于检测B细胞和T细胞的分裂。
用前准备：样本制成单细胞悬液；含0.1%BSA的PBS；5mM的CFSE贮存液；细胞培养基；
1．1重悬细胞于预温的PBS/0.1%BSA，浓度1×106/ml。
注意：为保证标记均匀，样本需为单细胞悬液。
细胞浓度为105~106,依标记后细胞的培养时间而定。
若需传代培养，浓度增加为1～5×107。
1．2每ml细胞中加入2ul CFSE贮存液，终浓度为10uM。
注意：最适工作浓度需优化，所以用前可用DMSO对贮存液进一步稀释，工作浓度的范围在0.5–25uM之间。
1．337℃孵育30min。
1．4加入5倍体积的冷培养基，终止染色。
1．5冰上孵育5min。
1．6离心沉淀细胞。
1．7用新鲜培养基洗3次。
1．8细胞置于条件合适的培养基或传代培养基中。
1．9收集细胞，进行其他染色。
1．10流式488nm激发光检测。
悬液中标记细胞的另一种方法
2．1离心收集细胞，在悬液中吹打均匀。
2．2用PBS或其他缓冲液稀释CFSE贮存液至工作浓度（0.5–25uM）。
2．3
2．437℃孵育细胞15min。
2．5沉淀，于新鲜预温的培养基中重悬。
2．6继续孵育30min，洗细胞。
贴壁细胞的标记
3．1细胞于培养皿中培养至所需密度。
3．2用PBS或其他缓冲液稀释CFSE贮存液至工作浓度（0.5–25uM）。
3．3移去培养皿中的培养基，加入预温至37℃ 含其他标记试剂的PBS。
3．437℃孵育细胞15min。
3．5将缓冲液换成新鲜培养基，37℃继续孵育30min。
在此过程中，CFSE醋酸盐水解。
（可选步骤）固定和破膜后进行其他标记
4．1固定前，用PBS或其他缓冲液洗细胞。
4．2标准的固定步骤，用含乙醛的固定剂，容易与蛋白－染料复合物的氨基发生交叉反应；可用3.7%的甲醛室温固定15min。
4．3固定后，用PBS冲洗。
4．4破膜试剂无特殊要求，如冰乙酮孵育10min。
破膜后，用PBS洗细胞。
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