基本描述

稳定性与储存	4℃保存。长期不使用可以-20℃保存。
英文名称	UltraBio™293 Transfection Reagent
储存温度	-20°C储存
运输条件	超低温冰袋运输
产品介绍	阿拉丁生产的UltraBio™293转染试剂(UltraBio™293 Transfection Reagent)是一种非常经济和高效的主要用于贴壁生长的HEK293、HEK293T、HEK293A、293FT等293系列细胞的基于新型阳离子脂质体的转染试剂。UltraBio™293转染试剂对HEK293细胞系列的转染效率约70%以上，同时其具有细胞毒性低、重复性好、操作简单、转染后无需更换培养基以及成本低等突出优点。贴壁细胞转染试剂的比较和选择请参考：http://www.aladdin-e.com。UltraBio™293转染试剂也能用于其它贴壁细胞的转染，但转染效率远低于阿拉丁生产的UltraBio™6000转染试剂和UltraBio™8000转染试剂。因此不太推荐使用UltraBio™293转染试剂用于293系列之外的细胞转染，仅当用于荧光素酶报告基因等检测灵敏度特别高的实验，并且用于一些比较容易转染的细胞时，才可以考虑使用UltraBio™293转染试剂。UltraBio™293转染试剂转染时血清的存在不会影响转染效率，这样可以减少避免很多阳离子脂质体因为需要在转染时去除血清而对细胞造成的损伤。UltraBio™293转染试剂转染质粒进入贴壁培养的HEK293等293系列细胞后，通常在24-48小时后达到比较理想的蛋白表达水平。UltraBio™293转染试剂的转染效率可以通过转染表达EGFP或其它荧光蛋白的质粒进行快速鉴定。UltraBio™293转染试剂的转染效果请参考图1。图1. UltraBio™293转染试剂转染的转染效果图。UltraBio™293转染试剂转染EGFP表达质粒至HEK293 (A, B)、HEK293T (C, D)和293FT (E, F)细胞的效果图。其中A、C、E为荧光照片；B、D、F为相应的明场照片。每毫升UltraBio™293转染试剂大约可以转染10厘米培养皿33个、6厘米培养皿100个、6孔板200个孔、12孔板500个孔、24孔板1000个孔、48孔板2000个孔、96孔板5000个孔。注意事项：使用高纯度的DNA有助于获得较高的转染效率。对于质粒，可以使用阿拉丁生产的质粒大量抽提试剂盒进行抽提，以保证可以获得较高的转染效率。转染前细胞必须处于良好的生长状态。对于非293系列细胞，推荐使用阿拉丁生产的Lipo6000?转染试剂或Lipo8000?转染试剂。对于悬浮培养的293系列细胞，推荐使用阿拉丁生产的Lipo293F?转染试剂。需自备不含抗生素的无血清培养液或Opti-MEM? Medium。Lipo293?转染试剂不能vortex或离心，可以用移液器轻轻吹打混匀或缓慢摇动混匀。Lipo293?转染试剂使用后请立即盖好盖子，避免长时间暴露在空气中，影响转染效率。本产品仅限于专业人员的科学研究用，不得用于临床诊断或治疗，不得用于食品或药品，不得存放于普通住宅内。为了您的安全和健康，请穿实验服并戴一次性手套操作。使用说明： 1.细胞培养(以六孔板为例，其它培养板或培养皿参考六孔板)：在转染前一天(18-24小时)把约20-60万细胞(具体的细胞数量据细胞类型、大小和细胞生长速度而定)培养到六孔板内，使第二天细胞能达到约60-70%。2.在进行下述转染步骤前，把培养有细胞的六孔板每孔换成2ml新鲜培养液(含有血清，不含抗生素)。可以使用含有血清并含有抗生素的新鲜培养液，但抗生素的存在对于有些细胞容易导致转染后出现一定的细胞毒性。3.参考下表，对于待转染的六孔板中每一个孔的细胞，取两个洁净无菌离心管，分别加入125μl不含抗生素和血清的DMEM培养液(高糖DMEM或低糖DMEM均可)或Opti-MEM® Medium，然后于其中一管加入2.5μg质粒DNA，并用枪轻轻吹打混匀；另一管加入5μlLipo293™转染试剂，用枪轻轻吹打混匀，请特别注意不可Vortex或离心。将含有DNA的培养液用枪轻轻加入含Lipo293™转染试剂的培养液中，轻轻颠倒离心管或者用枪轻轻吹打混匀，室温静置15分钟(室温存放6小时内稳定)。96-well48-well24-well12-well6-well 6cm dish10cm dishLipo293™转染试剂0.2μl0.5μl1μl2μl5μl 10μl30μl无血清培养液或Opti-MEM® Medium5μl12.5μl25μl50μl125μl 250μl750μlDNA100ng250ng500ng1μg2.5μg 5μg15μg无血清培养液或Opti-MEM® Medium5μl12.5μl25μl50μl125μl 250μl750μl单独稀释好的Lipo293™转染试剂和DNA，混匀并室温静止放置15分钟每孔加入的混合物的量10μl25μl50μl100μl250μl 500μl1500μl按照上述用量每孔均匀滴加Lipo293细胞转染试剂和DNA的混合物，继续培养48-72小时注1：对于六孔板中一个孔的细胞，Lipo293™转染试剂的用量可以在3-12.5μl内进行适当调节，DNA用量可以在1-4μg的范围内进行适当调节。质粒用量(μg)和Lipo293™(μl)的用量1:2比较常用。最佳的转染条件，因不同细胞类型和培养条件而定，可以在上述推荐范围内自行优化转染条件。注2：对于多个孔转染相同数量相同质粒的情况，可以把每个孔所需的Lipo293™转染试剂和DNA混合物分别配制，然后一起混合在同一个离心管内，后续混匀并孵育20分钟后，可以按照推荐用量滴加到细胞培养器皿内。注3：对于其它培养板或培养器皿，各种试剂的用量可以按照细胞培养面积按比例进行换算。如果转染RNA或寡核苷酸等可以参考转染DNA的条件进行。4.按照六孔板每孔250μl Lipo293™转染试剂-DNA混合物的用量，均匀滴加到整个孔内，随后轻轻混匀。5.继续培养约24-48小时后，即可用适当方式检测转染效果，例如荧光检测、Western、ELISA、报告基因等，或加入适当的筛选药物如G418等进行稳定细胞株的筛选。常见问题：1.转染效率低：a.优化质粒与Lipo293™转染试剂比例，适当加大质粒用量。b.应使用高纯度、无菌、无污染物的质粒进行转染，DNA纯度方面A260/A280比值要接近1.8，通常宜控制在1.8-1.9范围内，偏低则有可能有蛋白污染，偏高则有可能有RNA污染。可以使用阿拉丁生产的质粒大量抽提试剂盒进行抽提，以保证可以获得较高的转染效率。c.需用无抗生素和无血清培养液配制Lipo293™转染试剂和质粒的混合物。d.细胞转染时应状态良好，并且使用本产品时细胞密度达到60-70%时最适合进行转染，过稀或过密都可能影响转染效率，不同细胞的最佳转染密度需要自行摸索。e.转染后培养时间不足，而被误认为转染效率偏低。细胞转染后至显著表达所需培养时间通常为24-48小时。f.检查细胞是否支原体感染，支原体感染会影响细胞增殖，并很可能影响转染效率。g.如果没有检测到目的蛋白表达，应该仔细核对转染质粒的测序结果，确保测序结果和读码框完全正确。启动子、复制起始位点、质粒大小都会影响基因表达水平。2.出现一定的细胞毒性：a.转染前，细胞至少铺板18-24小时。 b.目的基因的表达蛋白有毒性。此时可以和空载质粒转染效果比较，以确定是否是目的基因蛋白表达对细胞产生毒性。如果确定有细胞毒性，可以考虑适当减少质粒用量，并按照比例减少Lipo293™转染试剂。c.检查是否转染时细胞密度太低。d.检查细胞是否有支原体等微生物污染。附录：常用多孔板和培养皿的尺寸、培养面积、细胞培养量和推荐的培养体积等相关数据表：Multiple Well Plates or DishesSingle Well Only for PlatesDiameter (Bottom, mm)Growth Area(cm2)Average Cell YieldTotal Well Volume (ml)WorkingVolume (ml)Recommended Volume (ml)6 well34.89.59.5 × 105 16.81.9-2.9212 well22.13.83.8 × 105 6.90.76-1.14124 well15.61.91.9 × 105 3.40.38-0.570.548 well11.00.959.5 × 104 1.60.19-0.2850.2596 well6.40.323.2 × 104 0.360.10-0.200.1384 well2.70.0565.6 × 103 0.1120.025-0.0500.0301536 well1.63 × 1.630.0252.5 × 103 0.01250.005-0.0100.0103.5 cm dish3499.0 × 105 NA1.8-2.726 cm dish52212.1 × 106 NA4.2-6.3510 cm dish8.4555.5 × 106 NA11-16.51215cm dish141521.5 × 107 NA30.4-45.63524.5cm dish22.4 × 22.45005.0 × 107 NA100-150120Diameter and growth area may vary depending on the manufacturer, and the listed sizes are from Corning. These wells or dishes are square. Aladdin's UltraBio™293 Transfection Reagent is an economical, cationic liposome-based formulation, designed specifically for highly efficient transfection of adherent cells, such as HEK293, HEK293T, HEK293A, 293FT, and other 293 cell lines.UltraBio™293 Transfection Reagent provides high transfection efficiency (over 70%) on HEK293 cell lines, with good reproducibility, simple operation, and extremely low cytotoxicity, and therefore changing culture medium is unnecessary after transfection. For more information about aladdin Transfection Reagents for transfecting adherent cells, please visit our website at http Precautions：* Use high-quality DNA to achieve higher transfection efficiency. We recommend isolating plasmid DNA using ’s Plasmid Maxi Preparation Kit (D0026).Make sure that the cells are actively dividing and reach the appropriate density at the time of transfection.To transfect adherent cells other than 293 cell lines, we recommend using Lipo6000™ Transfection Reagent (, C0526) or Lipo8000™ Transfection Reagent (, C0526). To transfect suspension 293 cell lines, the Lipo293F™ Transfection Reagent (, C0518) is recommended.Antibiotic-free and serum-free culture medium or Opti-MEM® Medium is required, but not supplied in the product.Mix Lipo293™ Transfection Reagent well gently prior to each use. Vortex or centrifuge should be avoided.Seal the Lipo293™ Transfection Reagent tightly after each use to avoid long time exposure to air that may reduce the transfection efficiency.This product is for R&D only. Not for drug, household, or other uses.For your safety and health, please wear a lab coat and disposable gloves during the operation. Instructions for Use： The following procedures are for the introduction of plasmid DNA into adherent 293 cell lines in 6-well plates. To transfect cells in other formats of cell culture vessels, please adjust the amount of Transfection Reagent proportionally as indicated in the table below.1. Seed 293 cell lines in 6-well culture plates: The day before transfection (18-24 hours in advance), seed 2-6×105 cells per well, depending on the cell type, cell size, and the growth rate of cells. Grow cells in appropriate conditions overnight to reach a cell density of 60-70% at the time of transfection.2. Before proceeding to the following transfection steps, replace per well with 2ml of fresh antibiotic culture medium containing serum. A culture medium containing both antibiotics and serum can also be used, but the presence of antibiotics may cause cytotoxicity after transfection. 3. Prepare Lipo293™ Transfection Reagent/DNA complexesa. Dilute 2.5µg of plasmid DNA in 125µl of antibiotic-free and serum-free DMEM culture medium (either high or low sugar DMEM) or Opti-MEM® Medium in a sterile tube. Mix gently. Vortex or centrifuge should be avoided.b. Dilute 5μl of Lipo293™ Transfection Reagent in 125µl of antibiotic-free and serum-free DMEM culture medium (either high or low sugar DMEM) or Opti-MEM® Medium in a sterile tube. Mix gently. Vortex or centrifuge should be avoided.c. Add the diluted DNA to the diluted transfection reagent. Mix gently. d. Incubate for 15 minutes at room temperature to allow the Lipo293™ Transfection Reagent/DNA complexes to form. The complexes are stable at room temperature for 6 hours.96-well48-well24-wel12-wel6-well6cm dish10cm dishLipo293™ Transfection Reagent0.2μl0.5μl1μl2μl5μl10μl30μlSerum-free or Opti-MEM® Medium5μl12.5μl25μl50μl125μl250μl750μlDNA100ng250ng500ng1μg2.5μg5μg15μgSerum-free or Opti-MEM® Medium5μl12.5μl25μl50μl125μl250μl750μlNote 1: The dose of Lipo293™ Transfection Reagent can be appropriately adjusted in the range of 3-12.5μl for each well of a 6-well plate. DNA dosage can be adjusted in the range of 1-4μg. A DNA (μg): Lipo293™(μl) ratio of 1:2 is generally used. The optimal transfection condition varies depending on the cell type and culture conditions and can be optimized within the range recommended above.Note 2: Prepare a master mix when transfecting multiple cell samples with the same DNA at the same amount, and then dispense the recommended amount to each well.Note 3: For other formats of cell culture vessels, the dose of each reagent can be scaled up or down accordingly based on the culture area of cell culture vessels. For the transfection of oligonucleotides or RNA, please refer to the protocol for the transfection of DNA.4. Add 250µl of Lipo293™ Transfection Reagent/DNA complexes drop-wise to each well of a 6-well plate. Gently shake the culture plate to evenly distribute the transfection complexes. 5. Continue to culture cells directly without changing the culture medium. After 24-48 hours of transfection, harvest cells and evaluate the transfection efficiency by appropriate methods such as fluorescence assay, Western Blot, ELISA, reporter gene, etc. To obtain cells with stable transfection, grow cells in a selective medium with antibiotics such as G418. FAQ:1. Low transfection efficiency:a. Optimize the ratio of plasmid DNA to Lipo293™ Transfection Reagent. Increase the plasmid amount if necessary.b. Use highly purified, sterile, and contaminant-free plasmid for transfection. High-purity plasmid should have an A260/A280 ratio ranging from 1.8 to 1.9. We recommend isolating plasmid DNA using Transfection Reagent/DNA complexes should be prepared in a culture medium without antibiotics and serum.d. Make sure that the cells to be transfected are in good condition. The cell density should be 60-70 % at the time of transfection. The optimal cell density for transfection should be determined for different cell types.e. Insufficient incubation time after transfection may result in low transfection efficiency. The recommended incubation time after transfection to yield high expression of target proteins is 24-48 hours.f. Regularly check cells for mycoplasma infection. Mycoplasma contamination detrimentally affects cell proliferation, and thereby the transformation efficiency.g. If the expression of the target protein is not detected, check the sequence of the plasmid to ensure the open reading frame is correct.2. Cellular toxicity occurs:a. Plat cells at least 18-24 hours in advance prior to transfection.b. To determine if the expressed target protein is toxic to cells, cells transfected with empty expression plasmid can be used as a negative control. If the target protein is indeed toxic to cells, reduction of plasmid amount as well as Lipo293™ Transfection Reagent amount accordingly can be considered. c. Check if the cell density is too low at the time of transfection.d. Check cell culture if there is any infection of mycoplasma or other microorganisms.Multiple Well Plates or DishesSingle Well Only for PlatesDiameter(Bottom, mm)Growth Area(cm2)AverageCell YieldTotal WellVolume (ml)WorkingVolume (ml)Recommended Volume (ml)6 well34.89.59.5 × 10516.81.9-2.9212 well22.13.83.8 × 1056.90.76-1.14124 well15.61.91.9× 105 3.40.38-0.570.548 well11.00.959.5× 104 1.60.19-0.2850.2596 well6.40.323.2 × 1040.360.10-0.200.1384 well2.70.0565.6 × 1030.1120.025-0.0500.0301536 well1.63 × 1.630.0252.5 × 1030.1250.005-0.0100.0103.5 cm dish3499.0 × 105NA1.8-2.726 cm dish52212.1 × 106NA4.2-6.3510 cm dish8.4555.5 × 106NA11-16.51215cm dish141521.5 × 107NA30.4-45.63524.5cm dish22.4 × 22.45005.0 × 107NA100-150120Diameter and growth area may vary depending on the manufacturer, and the listed sizes are from Corning. *These wells or dishes are square.

安全和危险性（GHS）

SDS英文版

质量标准

质量标准

质检证书（CoA,COO,BSE/TSE 和分析图谱)

C of A & Other Certificates(BSE/TSE, COO):

输入批号以搜索分析图谱:

溶液计算器

摩尔浓度计算器

计算溶液所需的质量、体积或浓度。

质量

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浓度

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体积

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分子量

g/mol

稀释计算器

计算配制储备溶液所需的稀释度。

浓度1（C1）

x

体积 1 (V1)

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浓度 2 (C2)

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体积 2 (V2)

复溶计算器

复溶计算器允许您快速计算试剂的体积，以复溶小瓶。只需输入试剂的质量和目标浓度，计算器将确定其余部分。

体积（添加到小瓶中）

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质量（小瓶）

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所需的复溶浓度

货号 (SKU)	包装规格	是否现货
T751606-500μl	500μL	期货
T751606-1.5ml	1.5ml	期货
T751606-5×1.5ml	5×1.5ml	期货

UltraBio™293转染试剂

库存信息

库存信息

库存信息

库存信息

基本描述

安全和危险性（GHS）

质量标准

质检证书（CoA,COO,BSE/TSE 和分析图谱)

溶液计算器

摩尔浓度计算器

稀释计算器

复溶计算器