PBS体内巨噬细胞清除剂 PBS空白脂质体对照|Control Liposomes(PBS)

PBS体内巨噬细胞清除剂 PBS空白脂质体对照|Control Liposomes(PBS)

产品说明书

FAQ

COA

已发表文献

氯膦酸二钠是一种特异的杀伤巨噬细胞的药物,可以通过诱导巨噬细胞凋亡来去除组织中的巨噬细胞。目前市场上理想的用于高效巨噬细胞耗竭的试剂是氯膦酸二钠脂质体,氯膦酸二钠分子包被在脂质体胶囊中,具有活性高、稳定性高及使用方便等特点。静脉注射0.2 mL 氯膦酸二钠脂质体24 h后脾脏(红髓巨噬细胞)中巨噬细胞清除率达到80%-90%。

该产品是不含氯膦酸二钠的脂质体,脂质体中仅含有PBS,可用于对照组实验,以了解巨噬细胞的消耗是否归因于注射氯膦酸二钠脂质体后产生的效果。

 

产品信息

货号

40338ES08 / 40338ES10

规格

5 mL / 10 mL

 

组分信息

组分名称

40338ES08

40338ES10

Control Liposomes ( PBS )

5 mL

10 mL

 

储存条件

2~8℃保存,有效期6个月

注意:

不能冻存!使用前一定要充分混匀并恢复到室温。

 

使用说明

该产品是作为对照组试验进行使用,具体使用剂量应与氯膦酸二钠脂质体(货号:40337)使用剂量一致。

 

注意事项

1. 为了您的安全和健康,请穿实验服并戴一次性手套操作。

2. 使用前一定要充分混匀并恢复到室温。

3. 本产品仅用于科研用途,禁止用于人身。

 

Ver.CN20230331

 

Q:体内巨噬细胞清除剂空白脂质体对照成分除了脂质体和pbs外还有其他成分吗

A: 没有

Q:请问买了巨噬细胞清除剂需要要买体内巨噬细胞清除剂空白脂质体对照吗?

A: 具体看实验设计,有的不需要

Q:体内巨噬细胞清除剂空白脂质体对照是聚乙二醇化脂质体吗?客户想要商用脂质体阿霉素类似的空白对照脂质体。

A:凑合可以两种产品的脂质体还是有点差别可以试试反正是对照脂质体的构成,每个厂家不一样好些客户就直接用PBS做对照,那个差的更远,但也是有参考意义

[1] Zhang Z, Chen C, Yang F, et al. Itaconate is a lysosomal inducer that promotes antibacterial innate immunity [published online ahead of print, 2022 May 25]. Mol Cell. 2022;S1097-2765(22)00443-9. doi:10.1016/j.molcel.2022.05.009(IF:17.970)
[2] Cai J, Peng J, Zang X, et al. Mammary Leukocyte-Assisted Nanoparticle Transport Enhances Targeted Milk Trace Mineral Delivery [published online ahead of print, 2022 Jun 30]. Adv Sci (Weinh). 2022;e2200841. doi:10.1002/advs.202200841(IF:17.521)
[3] Hu J, Deng F, Zhao B, et al. Lactobacillus murinus alleviate intestinal ischemia/reperfusion injury through promoting the release of interleukin-10 from M2 macrophages via Toll-like receptor 2 signaling. Microbiome. 2022;10(1):38. Published 2022 Mar 3. doi:10.1186/s40168-022-01227-w(IF:14.652)
[4] Sheng D, Ma W, Zhang R, et al. Ccl3 enhances docetaxel chemosensitivity in breast cancer by triggering proinflammatory macrophage polarization [published correction appears in J Immunother Cancer. 2022 Jun;10(6):]. J Immunother Cancer. 2022;10(5):e003793. doi:10.1136/jitc-2021-003793(IF:13.751)
[5] Zhang X, Hou L, Li F, et al. Piezo1-mediated mechanosensation in bone marrow macrophages promotes vascular niche regeneration after irradiation injury. Theranostics. 2022;12(4):1621-1638. Published 2022 Jan 16. doi:10.7150/thno.64963(IF:11.556)
[6] Sun Z, Huang W, Zheng Y, et al. Fpr2/CXCL1/2 Controls Rapid Neutrophil Infiltration to Inhibit Streptococcus agalactiae Infection. Front Immunol. 2021;12:786602. Published 2021 Nov 24. doi:10.3389/fimmu.2021.786602(IF:7.561)
[7] Zheng Y, Wang Z, Wei S, Liu Z, Chen G. Epigenetic silencing of chemokine CCL2 represses macrophage infiltration to potentiate tumor development in small cell lung cancer. Cancer Lett. 2021;499:148-163. doi:10.1016/j.canlet.2020.11.034(IF:7.360)
[8] Qian S, Han X, Sha X, et al. Aqueous Extract of Cimicifuga dahurica Reprogramming Macrophage Polarization by Activating TLR4-NF-κB Signaling Pathway. J Inflamm Res. 2022;15:1027-1046. Published 2022 Feb 15. doi:10.2147/JIR.S345497(IF:6.922)
[9] Wang X, Li W, Jiang H, et al. Zebrafish Xenograft Model for Studying Pancreatic Cancer-Instructed Innate Immune Microenvironment. Int J Mol Sci. 2022;23(12):6442. Published 2022 Jun 9. doi:10.3390/ijms23126442(IF:5.924)
[10] Ma Y, Liang Y, Wang N, et al. Avian Flavivirus Infection of Monocytes/Macrophages by Extensive Subversion of Host Antiviral Innate Immune Responses. J Virol. 2019;93(22):e00978-19. Published 2019 Oct 29. doi:10.1128/JVI.00978-19(IF:5.103)
[11] Jiang Q, Li W, Zhu X, et al. Estrogen receptor β alleviates inflammatory lesions in a rat model of inflammatory bowel disease via down-regulating P2X7R expression in macrophages. Int J Biochem Cell Biol. 2021;139:106068. doi:10.1016/j.biocel.2021.106068(IF:5.085)
[12] Shan K, Qiu J, Zhou R, et al. RNA-seq identifies long non-coding RNAs as potential therapeutic targets for human corneal endothelial dysfunction under oxidative stress. Exp Eye Res. 2021;213:108820. doi:10.1016/j.exer.2021.108820(IF:3.467)

氯膦酸二钠是一种特异的杀伤巨噬细胞的药物,可以通过诱导巨噬细胞凋亡来去除组织中的巨噬细胞。目前市场上理想的用于高效巨噬细胞耗竭的试剂是氯膦酸二钠脂质体,氯膦酸二钠分子包被在脂质体胶囊中,具有活性高、稳定性高及使用方便等特点。静脉注射0.2 mL 氯膦酸二钠脂质体24 h后脾脏(红髓巨噬细胞)中巨噬细胞清除率达到80%-90%。

该产品是不含氯膦酸二钠的脂质体,脂质体中仅含有PBS,可用于对照组实验,以了解巨噬细胞的消耗是否归因于注射氯膦酸二钠脂质体后产生的效果。

 

产品信息

货号

40338ES08 / 40338ES10

规格

5 mL / 10 mL

 

组分信息

组分名称

40338ES08

40338ES10

Control Liposomes ( PBS )

5 mL

10 mL

 

储存条件

2~8℃保存,有效期6个月

注意:

不能冻存!使用前一定要充分混匀并恢复到室温。

 

使用说明

该产品是作为对照组试验进行使用,具体使用剂量应与氯膦酸二钠脂质体(货号:40337)使用剂量一致。

 

注意事项

1. 为了您的安全和健康,请穿实验服并戴一次性手套操作。

2. 使用前一定要充分混匀并恢复到室温。

3. 本产品仅用于科研用途,禁止用于人身。

 

Ver.CN20230331

 

Q:体内巨噬细胞清除剂空白脂质体对照成分除了脂质体和pbs外还有其他成分吗

A: 没有

Q:请问买了巨噬细胞清除剂需要要买体内巨噬细胞清除剂空白脂质体对照吗?

A: 具体看实验设计,有的不需要

Q:体内巨噬细胞清除剂空白脂质体对照是聚乙二醇化脂质体吗?客户想要商用脂质体阿霉素类似的空白对照脂质体。

A:凑合可以两种产品的脂质体还是有点差别可以试试反正是对照脂质体的构成,每个厂家不一样好些客户就直接用PBS做对照,那个差的更远,但也是有参考意义

[1] Zhang Z, Chen C, Yang F, et al. Itaconate is a lysosomal inducer that promotes antibacterial innate immunity [published online ahead of print, 2022 May 25]. Mol Cell. 2022;S1097-2765(22)00443-9. doi:10.1016/j.molcel.2022.05.009(IF:17.970)
[2] Cai J, Peng J, Zang X, et al. Mammary Leukocyte-Assisted Nanoparticle Transport Enhances Targeted Milk Trace Mineral Delivery [published online ahead of print, 2022 Jun 30]. Adv Sci (Weinh). 2022;e2200841. doi:10.1002/advs.202200841(IF:17.521)
[3] Hu J, Deng F, Zhao B, et al. Lactobacillus murinus alleviate intestinal ischemia/reperfusion injury through promoting the release of interleukin-10 from M2 macrophages via Toll-like receptor 2 signaling. Microbiome. 2022;10(1):38. Published 2022 Mar 3. doi:10.1186/s40168-022-01227-w(IF:14.652)
[4] Sheng D, Ma W, Zhang R, et al. Ccl3 enhances docetaxel chemosensitivity in breast cancer by triggering proinflammatory macrophage polarization [published correction appears in J Immunother Cancer. 2022 Jun;10(6):]. J Immunother Cancer. 2022;10(5):e003793. doi:10.1136/jitc-2021-003793(IF:13.751)
[5] Zhang X, Hou L, Li F, et al. Piezo1-mediated mechanosensation in bone marrow macrophages promotes vascular niche regeneration after irradiation injury. Theranostics. 2022;12(4):1621-1638. Published 2022 Jan 16. doi:10.7150/thno.64963(IF:11.556)
[6] Sun Z, Huang W, Zheng Y, et al. Fpr2/CXCL1/2 Controls Rapid Neutrophil Infiltration to Inhibit Streptococcus agalactiae Infection. Front Immunol. 2021;12:786602. Published 2021 Nov 24. doi:10.3389/fimmu.2021.786602(IF:7.561)
[7] Zheng Y, Wang Z, Wei S, Liu Z, Chen G. Epigenetic silencing of chemokine CCL2 represses macrophage infiltration to potentiate tumor development in small cell lung cancer. Cancer Lett. 2021;499:148-163. doi:10.1016/j.canlet.2020.11.034(IF:7.360)
[8] Qian S, Han X, Sha X, et al. Aqueous Extract of Cimicifuga dahurica Reprogramming Macrophage Polarization by Activating TLR4-NF-κB Signaling Pathway. J Inflamm Res. 2022;15:1027-1046. Published 2022 Feb 15. doi:10.2147/JIR.S345497(IF:6.922)
[9] Wang X, Li W, Jiang H, et al. Zebrafish Xenograft Model for Studying Pancreatic Cancer-Instructed Innate Immune Microenvironment. Int J Mol Sci. 2022;23(12):6442. Published 2022 Jun 9. doi:10.3390/ijms23126442(IF:5.924)
[10] Ma Y, Liang Y, Wang N, et al. Avian Flavivirus Infection of Monocytes/Macrophages by Extensive Subversion of Host Antiviral Innate Immune Responses. J Virol. 2019;93(22):e00978-19. Published 2019 Oct 29. doi:10.1128/JVI.00978-19(IF:5.103)
[11] Jiang Q, Li W, Zhu X, et al. Estrogen receptor β alleviates inflammatory lesions in a rat model of inflammatory bowel disease via down-regulating P2X7R expression in macrophages. Int J Biochem Cell Biol. 2021;139:106068. doi:10.1016/j.biocel.2021.106068(IF:5.085)
[12] Shan K, Qiu J, Zhou R, et al. RNA-seq identifies long non-coding RNAs as potential therapeutic targets for human corneal endothelial dysfunction under oxidative stress. Exp Eye Res. 2021;213:108820. doi:10.1016/j.exer.2021.108820(IF:3.467)

脂质体核酸转染试剂 脂质体转染试剂|Hieff Trans™ Liposomal Transfection Reagent

脂质体核酸转染试剂 脂质体转染试剂|Hieff Trans™ Liposomal Transfection Reagent

产品说明书

FAQ

COA

已发表文献

产品描述

Hieff Trans®脂质体核酸转染试剂是一种多用途的脂质体转染试剂,适用于DNARNA和寡核苷酸的转染,对大多数真核细胞具有很高的转染效率。其独特的配方使其可直接加入培养基中,血清的存在不会影响转染效率,这样可以减少去除血清对细胞的损伤。转染后不需要除去核酸Hieff Trans®复合物或更换新鲜培养基,也可在46小时后除去。

Hieff Trans®脂质体核酸转染试剂以无菌的液体形式提供。通常情况下对于 24 孔板转染,每次用1.5 μL左右,则1 mL 约可做660 次转染;对于6孔板,每次用6 μL左右,则1 mL约可做160 次转染

 

运输与保存方法

冰袋(wet ice)运输。产品2-8ºC保存,一年有效。不可冷冻!

 

注意事项

1. Hieff Trans®脂质体核酸转染试剂要求细胞铺板密度较高,以60%-80%为佳,这有助于减少阳离子脂质体细胞毒性造成的影响具体铺板密度需要预实验摸索;如果你研究的基因要求比较长的表达时间,比如细胞周期相关基因,或者细胞表面蛋白,最好选择细胞铺板密度要求较低的转染试剂,不适合用脂质体核酸转染试剂。

2. Hieff Trans®脂质体核酸转染试剂可用于有血清培养基的转染,并且转染前后不需要换培养基。但是,制备转染复合物时要求用无血清培养基稀释DNA和转染试剂,因为血清会影响复合物的形成。另外,要检测所用的无血清培养基与脂质体核酸转染试剂的相容性,已知CD293, SFMII, VP-SFM就不相容。

3. 转染的时候培养基中不能添加抗生素。

4. 使用高纯度的DNARNA有助于获得较高的转染效率,质粒中的内毒素是转染的大敌。

5. 阳离子脂质体应该在2-8ºC保存,要注意避免多次反复长时间开盖,因为可能会导致脂质体氧化而影响转染效率。

6. 初次使用应优化DNA浓度和阳离子脂质体试剂量以得到最大的转染效率。DNA和转染试剂的比例,通常推荐是1:2-1:3,比如24孔板内接种0.5-2×105个细胞,使用0.5 µg DNA1-1.5 µL 转染试剂。通过调整DNA/Hieff Trans®脂质体核酸转染试剂比例优化转染效率,DNAμg: 试剂μL)比值在1:0.5-1:5

7. 本产品仅作科研用途!

 

操作流程(以24孔板为例,其他培养板加样体积请参考表一) 

【注】:转染试剂使用量受细胞类型及其他实验条件影响,建议初次使用时设置梯度进行优化最佳使用量。

贴壁细胞:转染前一天(20-24小时),胰酶消化细胞并计数,细胞铺板(不含抗生素),使其在转染时密度为70-95%0.5-2 × 105 cells/well for a 24-well plate)。

悬浮细胞:转染当天,配制DNA复合物之前,24孔板中细胞铺板,每500 µL生长培养基(不含抗生素)中加入4-8 × 105 cells

1. 按照以下体系配制DNA-Hieff Trans®脂质体核酸转染试剂复合物:

1)对于每孔细胞,使用50 μL无血清培养基(如OPTI-MEM 培养基)稀释0.5 μg DNA。混匀。

2)对于每孔细胞,使用50 μL无血清培养基(如OPTI-MEM 培养基)稀释0.6-2.5 μL Hieff Trans®脂质体核酸转染试剂。

 

Hieff Trans®脂质体核酸转染试剂稀释后室温孵育5 min(在30 min内同稀释的DNA 混合,保温时间过长会降低活性)

【注意】:即使脂质体核酸转染试剂使用OPTI-MEM 稀释,细胞也可以使用DMEM培养。如果DMEM作为脂质体核酸转染试剂的稀释液,必须在5 min内同稀释的DNA混合。

2. 混合稀释的DNA和稀释的脂质体核酸转染试剂(总体积100 µL),轻轻混匀,并在室温(15-25)孵育20 min,使得DNA-脂质体复合物形成。此时溶液可能会混浊,但不会影响转染。

【注意】DNA-脂质体复合物室温至少稳定保存5 h

3. 直接将100 µL DNA-Hieff Trans®复合物加入到细胞培养板每个孔中,摇动培养板,轻轻混匀。

【注意】:如果在无血清条件下转染,使用含血清的正常生长培养基进行细胞铺板。在加入复合物前移去生长培养基,替换为500 µL无血清培养基。

4. 375% CO2培养箱培养24-48 h,直至进行转基因表达分析,无需去掉复合物或更换培养基。然而,可能有必要在4-6 h后更换生长培养基,不会降低转染活性。

稳转细胞株:转染24 h后,按照1:10或更高比例在细胞中加入新鲜生长培养基,转染48 h后加入筛选培养基。

悬浮细胞株:在细胞中加入DNA-Hieff Trans®复合物后,如果需要可以4 h后加入PMA/PHA。对于Jurkat细胞,PHAPMA的终浓度分别为1 µg/mL50 ng/mL,可以提高CMV启动子活性和基因表达。对于K562细胞,只加入PMA足以提高启动子活性。

 

转染体系的调整

对于不同的细胞培养板,Hieff Trans®脂质体核酸转染试剂、DNA、细胞和培养基的使用量会有所不同,具体请参考下表(表一)。对于96 孔板培养,不需要提前一天进行细胞铺板,可以直接在平板中制备复合物,然后将细胞悬浮液加入到复合物就可以了,这样进一步减少了转染时间。这种改进步骤已经过293-H293-FCOS-7LCHO细胞的试验,同传统方法相比活性稍低。快捷的步骤和蛋白表达细胞系的高效转染使得脂质体核酸转染试剂非常适用于96 孔板的高通量转染,比如cDNA文库的筛选和蛋白瞬时表达。

表一 在不同的培养容器中转染,脂质体核酸转染试剂,核酸,细胞和培养基的用量

Culture vessel

Surf. area per well1

Shared reagents

DNA transfection

RNAi transfection

Vol. of plating medium

Vol. of dilution medium2

DNA

脂质体核酸转染试剂

RNA

脂质体核酸转染试剂

96-well

0.3 cm2

100 μL

2×25 μL

0.1 μg

0.2-0.5 μL

5 pmol

0.25 μL

24-well

2 cm2

500 μL

2×50 μL

0.5 μg

0.6-2.5 μL

20 pmol

1.0 μL

12-well

4 cm2

1 mL

2×100 μL

1 μg

2-4.5 μL

40 pmol

2.0 μL

6-well

10 cm2

2 mL

2×250 μL

2-4 μg

5-10 μL

100 pmol

5 μL

60-mm

20 cm2

5 mL

2×0.5 mL

4-8 μg

10-20 μL

200 pmol

10 μL

10-cm

60 cm2

15 mL

2×1.5 mL

12-24μg

30-60 μL

600 pmol

30 μL

1 不同厂商提供的细胞培养板表面积可能有所不同;

2 稀释DNARNAi所用的培养基体积。

【注】:该表使用量仅供参考,具体使用量还需根据细胞类型及其他实验条件进行优化。使用时DNAμg: Hieff Trans®μL)比值保持在1:0.5-1:5

 

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Polyethylenimine Linear(PEI) MW40000rapid lysis)线性PEI转染试剂(速溶型)MW40000

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HB220930

 

Q:脂质体转染的效率多少,毒性如何?

A:有些细胞如 293T293FT、Hela 等转染效率基本在85%以上;所有阳离子脂质体转染试剂对细胞都会存在一定的毒性,但是我们公司的转染试剂经过配方优化后其毒性大大降低,且转染效率也有进一步提升。

Q:转染试剂转染后需要换液吗?

A:对于换液可以区分两种情况;1、转染之前如果没有换液应在转染 6 小时左右后换液,以保证细胞生长所需营养,2、如果转染之前如果有换液,可以按照平时等到培养基出现营养不足时换液。

Q:转染试剂转单个质粒和多质粒共转的效率如何?

A:单转效率对于验证过的细胞效率都是很好,可以参考FAQ-验证过的细胞系,对于共转由于要涉及到质粒的混合比例和质粒与转染试剂的添加比例问题,因此具体的效率需要做相应的验证。

Q:转染试剂可以冻存吗?

A:不可以冻存,因为转染试剂是一种脂质体阳离子转染试剂,由于脂质体是不能在低温下冻存,因此转染试剂最好是 4 度储存,保持最好的转染效能。

Q:转染实验过程中是否需要更换成无血清培养基?

A:不需要,我们的转染试剂可以在含血清的介质中进行转染的过程。

Q:转染后需要进行终止反应吗?

A:不需要。脂质体复合物可以稳定存在 6 个小时。如果在进行转染前没有进行细胞换液,为了保证细胞正常生长所需的营养,需要在 4~6 小时后换用新的培养基。但如果转染之前已进行过换液则在脂质体转染后不需要进行再次换液。

Q:转染试剂毒性相比之前的批次大?

A:40802产品进行的工艺优化,纯度增高,相应的转染效率也随之变高,建议质粒与转染试剂的比例在1:2进行调整,一旦出现细胞死亡的现象,降低转染试剂比列。或者转染6h后进行换液。

Q: 它的大致成分和脂质体粒径,我们可以提供吗?

A: 提供不了粒径取决于客户的核酸和实验条件的 不是一个绝对值的。

Q:是稳转特制的转染试剂吗?若不是特制的转染试剂,那是特制的质粒才能进行稳转吗?

A:不是稳转特制的转染试剂,普通的转染试剂。质粒要求:稳转的质粒是普通的质粒,只是需要带有抗性,便于后期的筛选。建议:其中质粒转染受制于质粒大小、转染介质的限制,对很多细胞转染效率低,而且质粒整合入细胞基因组的效率极低,所以构建稳转株的成功率不高,请知悉,若做稳转细胞株,建议进行慢病毒包装(货号:41102)。

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产品描述

Hieff Trans®脂质体核酸转染试剂是一种多用途的脂质体转染试剂,适用于DNARNA和寡核苷酸的转染,对大多数真核细胞具有很高的转染效率。其独特的配方使其可直接加入培养基中,血清的存在不会影响转染效率,这样可以减少去除血清对细胞的损伤。转染后不需要除去核酸Hieff Trans®复合物或更换新鲜培养基,也可在46小时后除去。

Hieff Trans®脂质体核酸转染试剂以无菌的液体形式提供。通常情况下对于 24 孔板转染,每次用1.5 μL左右,则1 mL 约可做660 次转染;对于6孔板,每次用6 μL左右,则1 mL约可做160 次转染

 

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冰袋(wet ice)运输。产品2-8ºC保存,一年有效。不可冷冻!

 

注意事项

1. Hieff Trans®脂质体核酸转染试剂要求细胞铺板密度较高,以60%-80%为佳,这有助于减少阳离子脂质体细胞毒性造成的影响具体铺板密度需要预实验摸索;如果你研究的基因要求比较长的表达时间,比如细胞周期相关基因,或者细胞表面蛋白,最好选择细胞铺板密度要求较低的转染试剂,不适合用脂质体核酸转染试剂。

2. Hieff Trans®脂质体核酸转染试剂可用于有血清培养基的转染,并且转染前后不需要换培养基。但是,制备转染复合物时要求用无血清培养基稀释DNA和转染试剂,因为血清会影响复合物的形成。另外,要检测所用的无血清培养基与脂质体核酸转染试剂的相容性,已知CD293, SFMII, VP-SFM就不相容。

3. 转染的时候培养基中不能添加抗生素。

4. 使用高纯度的DNARNA有助于获得较高的转染效率,质粒中的内毒素是转染的大敌。

5. 阳离子脂质体应该在2-8ºC保存,要注意避免多次反复长时间开盖,因为可能会导致脂质体氧化而影响转染效率。

6. 初次使用应优化DNA浓度和阳离子脂质体试剂量以得到最大的转染效率。DNA和转染试剂的比例,通常推荐是1:2-1:3,比如24孔板内接种0.5-2×105个细胞,使用0.5 µg DNA1-1.5 µL 转染试剂。通过调整DNA/Hieff Trans®脂质体核酸转染试剂比例优化转染效率,DNAμg: 试剂μL)比值在1:0.5-1:5

7. 本产品仅作科研用途!

 

操作流程(以24孔板为例,其他培养板加样体积请参考表一) 

【注】:转染试剂使用量受细胞类型及其他实验条件影响,建议初次使用时设置梯度进行优化最佳使用量。

贴壁细胞:转染前一天(20-24小时),胰酶消化细胞并计数,细胞铺板(不含抗生素),使其在转染时密度为70-95%0.5-2 × 105 cells/well for a 24-well plate)。

悬浮细胞:转染当天,配制DNA复合物之前,24孔板中细胞铺板,每500 µL生长培养基(不含抗生素)中加入4-8 × 105 cells

1. 按照以下体系配制DNA-Hieff Trans®脂质体核酸转染试剂复合物:

1)对于每孔细胞,使用50 μL无血清培养基(如OPTI-MEM 培养基)稀释0.5 μg DNA。混匀。

2)对于每孔细胞,使用50 μL无血清培养基(如OPTI-MEM 培养基)稀释0.6-2.5 μL Hieff Trans®脂质体核酸转染试剂。

 

Hieff Trans®脂质体核酸转染试剂稀释后室温孵育5 min(在30 min内同稀释的DNA 混合,保温时间过长会降低活性)

【注意】:即使脂质体核酸转染试剂使用OPTI-MEM 稀释,细胞也可以使用DMEM培养。如果DMEM作为脂质体核酸转染试剂的稀释液,必须在5 min内同稀释的DNA混合。

2. 混合稀释的DNA和稀释的脂质体核酸转染试剂(总体积100 µL),轻轻混匀,并在室温(15-25)孵育20 min,使得DNA-脂质体复合物形成。此时溶液可能会混浊,但不会影响转染。

【注意】DNA-脂质体复合物室温至少稳定保存5 h

3. 直接将100 µL DNA-Hieff Trans®复合物加入到细胞培养板每个孔中,摇动培养板,轻轻混匀。

【注意】:如果在无血清条件下转染,使用含血清的正常生长培养基进行细胞铺板。在加入复合物前移去生长培养基,替换为500 µL无血清培养基。

4. 375% CO2培养箱培养24-48 h,直至进行转基因表达分析,无需去掉复合物或更换培养基。然而,可能有必要在4-6 h后更换生长培养基,不会降低转染活性。

稳转细胞株:转染24 h后,按照1:10或更高比例在细胞中加入新鲜生长培养基,转染48 h后加入筛选培养基。

悬浮细胞株:在细胞中加入DNA-Hieff Trans®复合物后,如果需要可以4 h后加入PMA/PHA。对于Jurkat细胞,PHAPMA的终浓度分别为1 µg/mL50 ng/mL,可以提高CMV启动子活性和基因表达。对于K562细胞,只加入PMA足以提高启动子活性。

 

转染体系的调整

对于不同的细胞培养板,Hieff Trans®脂质体核酸转染试剂、DNA、细胞和培养基的使用量会有所不同,具体请参考下表(表一)。对于96 孔板培养,不需要提前一天进行细胞铺板,可以直接在平板中制备复合物,然后将细胞悬浮液加入到复合物就可以了,这样进一步减少了转染时间。这种改进步骤已经过293-H293-FCOS-7LCHO细胞的试验,同传统方法相比活性稍低。快捷的步骤和蛋白表达细胞系的高效转染使得脂质体核酸转染试剂非常适用于96 孔板的高通量转染,比如cDNA文库的筛选和蛋白瞬时表达。

表一 在不同的培养容器中转染,脂质体核酸转染试剂,核酸,细胞和培养基的用量

Culture vessel

Surf. area per well1

Shared reagents

DNA transfection

RNAi transfection

Vol. of plating medium

Vol. of dilution medium2

DNA

脂质体核酸转染试剂

RNA

脂质体核酸转染试剂

96-well

0.3 cm2

100 μL

2×25 μL

0.1 μg

0.2-0.5 μL

5 pmol

0.25 μL

24-well

2 cm2

500 μL

2×50 μL

0.5 μg

0.6-2.5 μL

20 pmol

1.0 μL

12-well

4 cm2

1 mL

2×100 μL

1 μg

2-4.5 μL

40 pmol

2.0 μL

6-well

10 cm2

2 mL

2×250 μL

2-4 μg

5-10 μL

100 pmol

5 μL

60-mm

20 cm2

5 mL

2×0.5 mL

4-8 μg

10-20 μL

200 pmol

10 μL

10-cm

60 cm2

15 mL

2×1.5 mL

12-24μg

30-60 μL

600 pmol

30 μL

1 不同厂商提供的细胞培养板表面积可能有所不同;

2 稀释DNARNAi所用的培养基体积。

【注】:该表使用量仅供参考,具体使用量还需根据细胞类型及其他实验条件进行优化。使用时DNAμg: Hieff Trans®μL)比值保持在1:0.5-1:5

 

相关产品

名称

货号

规格

价格(元)

Calcium Phosphate Cell Transfection Kit 磷酸钙法细胞转染试剂

40803ES70

200 T

625.00

Polybrene (hexadimethrine bromide) 聚凝胺(10 mg/ml

40804ES76

500 μL

180.00

40804ES86

5×500 μL

500.00

Hieff Trans® Suspension Cell-Free Liposomal Transfection Reagent 悬浮细胞专用脂质体核酸转染试剂

40805ES02

0.5 mL

948.00

40805ES03

1.0 mL

1678.00

40805ES08

5×1 mL

5268.00

Hieff Trans® in vitro siRNA/miRNA Transfection Reagent siRNA/miRNA体外转染试剂

40806ES02

0.5 mL

1472.00

40806ES03

1.0 mL

2572.00

Polyethylenimine Linear(PEI) MW25000 线性PEI转染试剂MW25000

40815ES03

1 g

1855.00

40815ES08

5×1 g

7255.00

Polyethylenimine Linear(PEI) MW40000rapid lysis)线性PEI转染试剂(速溶型)MW40000

40816ES02

100 mg

655.00

40816ES03

1 g

1855.00

HB220930

 

Q:脂质体转染的效率多少,毒性如何?

A:有些细胞如 293T293FT、Hela 等转染效率基本在85%以上;所有阳离子脂质体转染试剂对细胞都会存在一定的毒性,但是我们公司的转染试剂经过配方优化后其毒性大大降低,且转染效率也有进一步提升。

Q:转染试剂转染后需要换液吗?

A:对于换液可以区分两种情况;1、转染之前如果没有换液应在转染 6 小时左右后换液,以保证细胞生长所需营养,2、如果转染之前如果有换液,可以按照平时等到培养基出现营养不足时换液。

Q:转染试剂转单个质粒和多质粒共转的效率如何?

A:单转效率对于验证过的细胞效率都是很好,可以参考FAQ-验证过的细胞系,对于共转由于要涉及到质粒的混合比例和质粒与转染试剂的添加比例问题,因此具体的效率需要做相应的验证。

Q:转染试剂可以冻存吗?

A:不可以冻存,因为转染试剂是一种脂质体阳离子转染试剂,由于脂质体是不能在低温下冻存,因此转染试剂最好是 4 度储存,保持最好的转染效能。

Q:转染实验过程中是否需要更换成无血清培养基?

A:不需要,我们的转染试剂可以在含血清的介质中进行转染的过程。

Q:转染后需要进行终止反应吗?

A:不需要。脂质体复合物可以稳定存在 6 个小时。如果在进行转染前没有进行细胞换液,为了保证细胞正常生长所需的营养,需要在 4~6 小时后换用新的培养基。但如果转染之前已进行过换液则在脂质体转染后不需要进行再次换液。

Q:转染试剂毒性相比之前的批次大?

A:40802产品进行的工艺优化,纯度增高,相应的转染效率也随之变高,建议质粒与转染试剂的比例在1:2进行调整,一旦出现细胞死亡的现象,降低转染试剂比列。或者转染6h后进行换液。

Q: 它的大致成分和脂质体粒径,我们可以提供吗?

A: 提供不了粒径取决于客户的核酸和实验条件的 不是一个绝对值的。

Q:是稳转特制的转染试剂吗?若不是特制的转染试剂,那是特制的质粒才能进行稳转吗?

A:不是稳转特制的转染试剂,普通的转染试剂。质粒要求:稳转的质粒是普通的质粒,只是需要带有抗性,便于后期的筛选。建议:其中质粒转染受制于质粒大小、转染介质的限制,对很多细胞转染效率低,而且质粒整合入细胞基因组的效率极低,所以构建稳转株的成功率不高,请知悉,若做稳转细胞株,建议进行慢病毒包装(货号:41102)。

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integralmolecular 脂质体技术

Integral Molecular是一家研究驱动的生物技术公司,为未被开发的膜蛋白靶标(包括GPCR,离子通道,转运蛋白和病毒包膜)创造创新技术和治疗性抗体。整合膜蛋白包括关键的治疗靶,但由于其结构复杂性和不能在构象完整状态下从脂质膜中除去而难以研究。在Integral Molecular,我们开发创新技术和产品,以满足这些苛刻但重要的目标。

脂质体技术

病毒样颗粒上的高浓度膜蛋白,用于药物和抗体的发现。

为什么我需要Lipoparticles用于抗体免疫和筛查?

脂质颗粒是病毒样颗粒(VLP),其天然构象含有高浓度的特异性膜蛋白。脂质颗粒直接从细胞表面捕获构象完整的膜蛋白,使得这些复合蛋白能够作为可溶性蛋白进行操作,用于抗体免疫和筛选。

Lipoparticle

 

 

为什么Lipoparticles是有效的抗原?

  • Lipoparticles在其天然细胞膜中显示正确折叠的膜蛋白。这导致引发和筛选构象抗体的能力。
  • 与细胞相比,脂质颗粒含有10-100倍浓度的膜蛋白(~50-200pmol / mg),导致强烈的免疫应答和抗体筛选的更好成功。

Lipoparticle应用包括:

  • 免疫接种
  • 噬菌体/酵母展示
  • 通过ELISA筛选抗体
  • 生物传感器的动力学分析
  • 放射性配体和荧光结合试验

 

定制脂质颗粒

定制脂质颗粒以包含任何选择的膜蛋白。使用质量控制指标评估每批脂质颗粒,以确保均一性,纯度和目标蛋白质完整性。数百种膜蛋白靶标已成功掺入脂质颗粒中。对于特定应用,可以用生物素或荧光修饰脂质颗粒。

ReadyReceptor Lipoparticles

预先验证的Lipoparticles含有优化的,高度表达的膜蛋白,可用于快速递送。ReadyReceptor Lipoparticles使用Custom Lipoparticle生产中使用的所有严格质量指标生成和验证。

GPCR

 

      Receptor

Description

Catalog No.

       A2AR

Alpha Adrenergic

RR-0910

       B2AR

Beta Andrenergic

RR-0574

      CR

Complement

RR-0362

       CCR1

Chemokine

RR-0968

        CCR2b

Chemokine

RR-0125

        hCCR5

Chemokine

RR-0190

        mCCR5

Chemokine

RR-0835

      rhCCR5

Chemokine

RR-0895

      CCR6

Chemokine

RR-0822

     CCR10

Chemokine

RR-0193

     CXCR2

Chemokine

RR-0943

     CXCR3

Chemokine

RR-0132

     CXCR4

Chemokine

RR-0830

     CXCR7

Chemokine

RR-0656

     mCXCR4

Chemokine

RR-2194

     GLP-1R

Glucagon-like peptide-1 Receptor

RR-1090

      GCGR

Glucagon Receptor

RR-0999

      GAL1 Receptor

Neuropeptide Receptor

RR-1067

      5-HT2C

Serotonin Receptor

RR-1498

       FZD4

Wnt Pathway

RR-1302

 

 

     Ion Channel

      Receptor

Description

Catalog No.

      HV1

Voltage Gated H+

RR-0678

      TRPC4

Nonspecific Cation Channel

RR-1054

       TRPM4

Nonspecific Cation Channel

RR-1333

       K2P1.1

Potassium Channel

RR-1225

       K2P4.1

Potassium Channel

RR-1331

       P2X2

ATP Purinoceptor

RR-1181

 

 

      RTK

     Receptor

Description

Catalog No.

      ALK2

Activin Receptor

RR-0850

 

      1-TM

    Receptor

Description

Catalog No.

     ACVR1

Activin Receptor, Type 1

RR-0850

     CD19

B-Cell Activation

RR-0553

     CD27

B-Cell Activation

RR-0655

    DC-SIGN

Lectin

RR-0057

    DC-SIGNR

Lectin

RR-0056

    LRP6

Wnt Pathway

RR-0853

    hTL1A

TNF Ligand

RR-0819

    mTL1A

TNF Ligand

RR-0817

    DR5

TNF-family Receptor

RR-0818

    ST2L

IL-1 Receptor Family

RR-0653

 

    4-TM

    Receptor

Description

Catalog No.

     CD81

Hepatitis C Attachment

RR-0341

    CD20

B-Cell Activation

RR-1179

    Claudin-1

Tight Junction

RR-0825

    Claudin-3

Tight Junction

RR-0733

    Claudin-4

Tight Junction

RR-0686

 

    Viral Protein

    Receptor

Description

Catalog No. 

    RSV-F

Respiratory Syncytial Virus

RR-0891

      Influenza M2           A/Udorn/307/1972

Ion Channel

RR-0929

    Influenza M2     A/HK/156/1997

Ion Channel

RR-0179

  Chikungunya Virus

Viral Envelope

RR-0945

    Ross River Virus

Viral Envelope

RR-1136

 

    Transporter

     Receptor

Description

Catalog No. 

     GLUT1

Glucose Transporter

RR-0777

 

 

Sample Kits

Lipoparticle sample kits containing CXCR4, CCR5 or CD20 are available for overnight delivery to enable application optimization and small-scale testing.

样品套件

含有CXCR4,CCR5或CD20的Lipoparticle样品试剂盒可用于隔夜输送,以实现应用优化和小规模测试。

 

Sample Kit

Class

Cat. No

Size

CXCR4

GPCR

LEV-101, LEV-101B

400 U

CCR5

GPCR

LEV-102, LEV-102B

400 U

CD20

GPCR

LEV-103, LEV-103B

400 U

脂质颗粒可以用(B)生物素或(F)荧光修饰。

 

 

上海金畔生物科技有限公司是实验试剂一站式采购服务商

1:强大的进口辐射能力,血清、抗体、耗材、大部分限制进口品等。

2:产品种类齐全,经营超过700多个品牌,基本涵盖所有生物实验试剂耗材。

3:提供加急服务,货品一般1-2周到货。

4:富有竞争力的价格优势,绝大部分价格有优势。

5:多年积累良好的信誉,大部分客户提供货到付款服务。客户包括清华、北大、交大、复旦、中山等100多所高校,ROCHE,阿斯利康、国药、fisher等药企。

6:我们还是Santa,Advanced Biotechnologies Inc,Athens Research & Technology,bangs,BBInternational,crystalchem,dianova,FD Neurotechnologies,Inc. FormuMax Scientific,Inc, Genebridege, Glycotope Biotechnology GmbH; iduron,Innovative Research of America, Ludger, neuroprobe,omicronbio, Polysciences,prospecbi, QA-BIO,quickzyme,RESEARCH DIETS,INC,sterlitech;sysy,TriLink BioTechnologies,Inc;worthington-biochem,zyagen等几十家国外公司代理。

7:我们还是invitrogen,qiagen,MiraiBioam,sigma;neb,roche,merck, rnd,BD, GE,pierce,BioLegend等*批发,欢迎合作。

 

clodronateliposomes脂质体使用说明书

clodronateliposomes脂质体使用说明书

 

产品描述

磷脂双层由磷脂酰胆碱和胆固醇组成。将所有脂质体制剂悬浮在无菌磷酸盐缓冲盐水(PBS)中:

– 10 mM的Na2HPO4;
– 10毫米NaH2PO4;
– 140毫米氯化钠。

我们所有的脂质体均带负电荷,因此不会一起阻塞。悬浮液中的脂质体没有大小,这意味着它也包含越来越大的脂质体(大3微米)。平均而言,脂质体的大小为1.7微米。

氯膦酸盐脂质体:人工制备的脂质囊泡的悬浮液,包裹了氯膦酸盐。悬浮液中氯膦酸盐的浓度约为。5毫克/毫升 氯膦酸盐以CH2Na2Cl2O6P2·5 H2O的形式封装在脂质体囊泡中。

PBS脂质体:人工制备的脂质囊泡的悬浮液,其中包裹了PBS水溶液。这些不含氯膦酸盐,可用于对照实验。

荧光DiI脂质体:人工制备的脂质囊泡的悬浮液,其中包封了PBS水溶液,并用荧光染料DiI标记。它们不含氯膦酸盐,可用于研究通过特定给药途径注射的脂质体是否能够到达要研究的巨噬细胞。

 

储存和使用说明

到达脂质体后,应将其储存在4 – 8ºC(或39 – 47ºF)之间。脂质体悬浮液不得冷冻,也不要暴露于高温下。这可能会对磷脂双层造成干扰,可能导致氯膦酸盐从脂质体中泄漏出来。

给药前,首先使脂质体达到室温,然后轻轻摇动或搅拌悬浮液。一段时间后,脂质体易于沉淀,导致小瓶中的分布不均匀。当注射花费太多时间时,脂质体甚至可能在注射器中沉淀。如果使用同一注射器注射多只动物,可能会导致剂量不同。

不建议稀释悬浮液,但如有必要,请使用PBS或盐水。

我们建议客户在发货后16周内使用脂质体制剂。强烈建议不要在到期日期之后使用。在此期间之后,污染的风险会增加,并且可能会发生轻微的功能丧失。

作用机理

巨噬细胞在免疫和非免疫防御机制中起重要作用。它们是抵御渗透到脊椎动物体内的细菌,病毒和其他形式的微生物污染的首道防线。巨噬细胞是大型细胞,几乎存在于所有身体组织中,它们可以具有不同的形式和名称(例如库普弗细胞,肺泡巨噬细胞,小胶质细胞,破骨细胞,红髓巨噬细胞)。巨噬细胞“清除”,它们摄入并消化可能是潜在病原体的所有异物,微生物,癌细胞和细胞碎片。该过程称为吞噬作用。巨噬细胞主要通过可溶性分子如细胞因子和趋化因子的介导来进一步调节许多非吞噬细胞的功能。他们参与先天免疫,

脂质体是人工制备的球体,由同心磷脂双层组成。当磷脂分散在水中时,亲水性头部将构成脂质体的两个外部部分,而疏水性脂肪酸基团将构成内部部分(见图1)。水室分隔双层,亲水分子可以溶解在其中,形成脂质体包封的分子。氯膦酸盐(二氯亚甲基双膦酸酯或Cl2MBP)是一种亲水性分子,可以封装在磷脂双层中。游离氯膦酸盐不易穿过细胞膜,并通过肾脏系统从循环中迅速清除(即数分钟内)。但是,氯膦酸盐脂质体包埋在脂质体中时,会被巨噬细胞吞噬,无法逃脱(见图2)。磷脂双层被溶酶体磷脂酶消化,而氯膦酸盐未被消化并且保留在巨噬细胞中。巨噬细胞摄入的磷脂双层和脂质体越多,在巨噬细胞内积累的氯膦酸盐越多。超过一定的细胞内浓度,氯膦酸盐将通过启动程序性细胞死亡(即凋亡)来消除巨噬细胞。

图1.氯膦酸盐脂质体的示意图。囊泡由同心的磷脂双层构成,被水性隔室隔开。双层由亲水和疏水基团组成。将氯膦酸盐(在此以黑色方块表示)溶解在水溶液中并封装在脂质体内。

 

图2.巨噬细胞对氯膦酸盐脂质体的摄取和消化示意图。巨噬细胞通过胞吞作用摄入氯膦酸盐脂质体,然后与含有磷脂酶的溶酶体(L)融合(箭头)。磷脂酶破坏的磷脂双层越多,在巨噬细胞内释放的氯膦酸盐(黑色方块)越多。巨噬细胞终通过凋亡被杀死。(N =巨噬细胞核)。

 
因此,氯膦酸盐脂质体可用于通过消耗巨噬细胞来研究巨噬细胞功能。例如,它们可以应用于各种自身免疫性疾病,移植,神经系统疾病和基因治疗的模型。氯膦酸盐脂质体只有在可以达到的情况下才能耗尽巨噬细胞。一些组织可以形成脂质体的屏障。通过选择氯膦酸盐脂质体的正确给药途径,可以耗尽特定的器官或组织的巨噬细胞。有关管理协议的更多信息,请单击此处。

PBS脂质体主要用于对照实验。但是,这些也可以在某些时间段内通过饱和来阻止吞噬作用。因此,PBS脂质体不能代表正常健康的,未阻断的,未抑制的和未活化的巨噬细胞的对照实验。当比较氯膦酸盐脂质体和PBS脂质体的作用时,其作用因此可能小于预期。

有关更多信息,请参见:

– Van Rooijen,N.和Sanders,A.(1994)。脂质体介导的巨噬细胞耗竭:作用机理,脂质体制备和应用。免疫学杂志,174(1-2),83-93。
– Van Rooijen,N.,Sanders,A.和van den Berg,TK(1996)。脂质体介导的氯膦酸盐和丙am的细胞内递送诱导的巨噬细胞凋亡。免疫学杂志,193(1),93-99。

 

encapsula 荧光脂质体

encapsula 荧光脂质体(Fluoroliposome®)

 

  • 荧光脂质体 – 用于追踪和检测的脂质体
    • 含有亲脂示踪剂的脂质体
      • 含有亲脂示踪剂的负电荷荧光脂质体
        • 磷脂酰甘油脂质体
          • 基于PG的Fluoroliposome®用于跟踪和检测
        • 磷脂酰丝氨酸脂质体
          • 基于PS的Fluoroliposome®用于跟踪和检测
      • 含有亲脂示踪剂的中性荧光脂质体
        • 用于追踪和检测的中性Fluoroliposome®
      • 含有亲脂示踪剂的带正电荷的荧光脂质体
        • 基于DOTAP的Fluoroliposome®用于跟踪和检测
    • 表面活性脂质体用于活性染料的共轭
      • 负电荷荧光脂质体用于活性染料的共轭
        • 磷脂酰甘油脂质体
          • 基于PG的Fluoroliposome®-DBCO用于跟踪和检测
          • 基于PG的Fluoroliposome®-Azide用于跟踪和检测
        • 磷脂酰丝氨酸脂质体
          • 基于PS的Fluoroliposome®-DBCO用于跟踪和检测
          • 基于PS的Fluoroliposome®-Azide用于跟踪和检测
      • 中性荧光脂质体用于活性染料的共轭
        • 用于追踪和检测的中性荧光脂质体 – 叠氮化物
        • 中性Fluoroliposome®-DBCO用于跟踪和检测
      • 带正电荷的荧光脂质体用于活性染料的共轭
        • 基于DOTAP的Fluoroliposome®-DBCO用于跟踪和检测
        • 基于DOTAP的Fluoroliposome®-Azide用于跟踪和检测
  • 荧光脂质体 – 用于融合实验的脂质体
    • 脂质体共包封NBD /罗丹明
      • 负电荷荧光脂质体共包封NBD /罗丹明
        • 磷脂酰甘油脂质体
          • DOPC:DOPG脂质体共包封NBD /罗丹明
          • POPC:Chol:POPG脂质体共包封NBD /罗丹明
          • POPE:POPG脂质体共包封NBD /罗丹明
        • 磷脂酰丝氨酸脂质体
          • DOPC:DOPS脂质体共包封NBD /罗丹明
          • POPE:POPS:POPC脂质体共包封NBD /罗丹明
      • 中性荧光脂质体共包封NBD /罗丹明
        • DOPC脂质体共包封NBD /罗丹明
      • 带正电荷的荧光脂质体共包封NBD /罗丹明
        • DOTAP脂质体共包封NBD /罗丹明
    • 包封十八烷基罗丹明B的脂质体
      • 含有十八烷基罗丹明B的负电荷荧光脂质体
        • 磷脂酰甘油脂质体
          • POPC:Chol:含有十八烷基罗丹明B自猝灭浓度的POPG脂质体
          • DOPC:含有十八烷基罗丹明B自猝灭浓度的DOPG脂质体
          • POPE:含有十八烷基罗丹明B自猝灭浓度的POPG脂质体
        • 磷脂酰丝氨酸脂质体
          • DOPC:含有十八烷基罗丹明B自猝灭浓度的DOPS脂质体
          • POPE:POPS:含有十八烷基罗丹明B自猝灭浓度的POPC脂质体
      • 包封十八烷基罗丹明B的中性荧光脂质体
        • 含有十八烷基罗丹明B自猝灭浓度的DOPC脂质体
      • 包封十八烷基罗丹明B的带正电荷的荧光脂质体
        • DOTAP脂质体含有十八烷基罗丹明B的自猝灭浓度
  • 荧光脂质体 – 用于孔形成实验的脂质体
    • 含有各种大小的葡聚糖-FITC染料的脂质体
      • 含有不同大小的葡聚糖-FITC染料的负电荷荧光脂质体
        • 磷脂酰甘油脂质体
          • POPE:含有葡聚糖-FITC染料的POPG脂质体
          • POPC:Chol:含有葡聚糖-FITC染料的POPG脂质体
          • DOPC:含有葡聚糖-FITC染料的DOPG脂质体
        • 磷脂酰丝氨酸脂质体
          • POPE:POPS:含有葡聚糖-FITC染料的POPC脂质体
          • DOPC:含有葡聚糖-FITC染料的DOPS脂质体
      • 含有不同大小的葡聚糖-FITC染料的中性荧光脂质体
        • 含有葡聚糖-FITC染料的DOPC脂质体
      • 含有不同大小的葡聚糖-FITC的带正电荷的荧光脂质体
        • 含有葡聚糖-FITC染料的DOTAP脂质体
  • 荧光脂质体 – 用于巨噬细胞摄取的脂质体
    • 标准荧光控制
      • Fluoroliposome®-DIA
      • Fluoroliposome®结果来自
      • Fluoroliposome®-的DiI
      • Fluoroliposome®-DIO
      • Fluoroliposome®-DIR
      • 三色Fluoroliposome®试剂盒
      • 五色Fluoroliposome®试剂盒
    • 甘露糖化荧光控制
      • M-Fluoroliposome®-DIA
      • M-Fluoroliposome®-DiD的
      • M-Fluoroliposome®-的DiI
      • M-Fluoroliposome®-DIO
      • M-Fluoroliposome®-DIR
      • 三色甘露糖基Fluoroliposome®试剂盒
      • 五色甘露糖化Fluoroliposome®试剂盒
  • 荧光脂质体 – 表面反应性脂质体(IMMUNOFLUOR™)
    • 胺反应性荧光脂质体
      • ImmunoFluor™-NHS(聚乙二醇化)(插入后)
      • ImmunoFluor™-Glutaryl(非PEG化)
      • ImmunoFluor™ – 琥珀酰(非聚乙二醇化)
      • ImmunoFluor™-Dodecanyl(非PEG化)
      • ImmunoFluor™ – 琥珀酰(PEG化)
      • ImmunoFluor™-Cyanur(聚乙二醇化)
      • ImmunoFluor™ – 羧酸(聚乙二醇化)
    • 生物素化的荧光脂质体
      • ImmunoFluor™ – 生物素基帽(非PEG化)
      • ImmunoFluor™-Biotin(非PEG化)
      • ImmunoFluor™-Biotin(聚乙二醇化)
    • 羧酸反应荧光脂质体
      • ImmunoFluor™-Amine(聚乙二醇化)
      • ImmunoFluor™ – 十二烷基胺(非PEG化)
      • ImmunoFluor™-Caproylamine(非PEG化)
    • DBCO或叠氮化物反应荧光脂质体(点击化学)
      • ImmunoFluor™-Azide(非PEG化)
      • ImmunoFluor™-DBCO(非PEG化)
      • ImmunoFluor™-Azide(聚乙二醇化)
      • ImmunoFluor™-DBCO(聚乙二醇化)
    • 叶酸荧光脂质体
      • ImmunoFluor™-Folate(聚乙二醇化)
    • 巯基反应性荧光脂质体
      • ImmunoFluor™-PDP(非PEG化)
      • ImmunoFluor™-PDP(聚乙二醇化)
      • ImmunoFluor™ – 马来酰亚胺(聚乙二醇化)(插入后)

clodronateliposomes氯膦酸二钠脂质体*


Clodronate Liposomes体内巨噬细胞清除试剂

去除巨噬细胞是研究巨噬细胞功能的重要方法。巨噬细胞可吞噬 clodronate Liposome, 在溶酶体的作用下,clodronate 可释放出来,当其达到一定浓度时,可引起巨噬细胞的凋亡, 从而去除巨噬细胞。clodronate Liposome 可局部注射去除注射部位临近的巨噬细胞,如肺、 膝关节、肿瘤局部和脑等;也可尾静脉或腹腔注射去除循环途径中的单核巨噬细胞,如肝脏 和脾脏中的巨噬细胞。

1. 脂质体的准备 可以自己制备或者购买商品话的脂质体 荷兰clodronaiposomes

2. 脂质体的体内注射 隔日尾静脉注射 200ul clodronate liposome,根据实验要求制定注射日程。通过免疫组化 或流式检测巨噬细胞的去除效率。

尾静脉注射方法: 尾静脉注射方法:

 1. 将小鼠放在倒扣的饭盒内,从孔口拉出尾巴,小鼠的尾部有2条动脉和3条静脉,2条动脉分别在尾部的背侧面和腹侧面,3 条静脉呈品字型分布,一般采用左右两侧的静脉;

2. 将纱布浸泡于约 70 度的温水中,拿出后包裹尾巴,以达到使尾部血管扩张及软化表皮 角质的目的;

3. 行尾部静脉注射时,以左手拇指和食指前后摁住鼠尾,留出约 2cm 一段,使皮肤紧张, 静脉更为充盈,右手持 1ml 针头注射器,使针头与静脉平行(小于 30°角),从尾巴的下 1/4 处进针,开始注入药物时应缓慢,仔细观察,如果无阻力,无白色皮丘出现,说明 已刺入血管,可注入药物。

4. 有的实验需连日反复尾静脉注射给药,注射部位应尽可能从尾端开始,按次序向尾根部 移动,更换血管位置注射给药。拔出针头后,用棉球按住注射部位轻压 1-2min,止血。 

FAQ:

Q:收到Clodronate liposomes后该如何操作?

A:收到氯磷酸脂质体后,如果不能马上使用,需要放置在4℃冰箱。禁止冻存!使用时作为原液使用,不能进行稀释。脂质体很容易发生沉淀,建议使用之前轻轻混匀,另外4℃的氯磷酸脂质体不能马上使用,需要恢复到室温才能进行注射。

Q: Clodronate liposomes能否用于体外巨噬细胞清除?

A:产品可以用于体外巨噬细胞的清除,但是这种方法比较适合体内实验。主要是由于在体外培养过程中,从死亡细胞释放的氯磷酸或者是从脂质体中“渗漏”出来氯磷酸,会一直存在于培养基中,但是在体内,氯磷酸的半衰期很短,很快就会被肾脏清除。虽然游离的氯磷酸不会进入细胞也不会进入脂质体,但是一旦他们在培养基中积累就会慢慢地进入细胞中。

Q:在静脉注射Clodronate liposomes后动物很快就死亡了,是什么原因?

A:一方面有可能是动物注射了不均一的脂质体悬液。建议在使用之前轻轻摇晃混匀。脂质体在体外一段时间后会发生沉降。如果在很长一段时间内需要一次注射多只动物,那么脂质体便会在注射器中发生沉淀,形成不均一悬液,这样一来,*只注射的计量与zui后一只注射的计量就会发生明显的变化。另外一方面有可能是脂质体刚从冰箱取出后,没有恢复到室温。

Q:动物在注射Clodronate liposomes后几天就死亡了?

A:这个可能是受到细菌的污染,体内清除巨噬细胞后会增加例如病毒粒子,细菌或者酵母感染的几率。

Q:静脉注射Clodronate liposomes后大鼠脾脏/肝脏中的ED1+细胞并没有*消失?

A:大鼠中成熟的巨噬细胞呈现ED1和ED2双阳性,但是有一些没有吞噬功能的或者吞噬作用小的前体细胞也是ED1+,但是呈现ED2-。因此,所有ED2+的细胞都可以被*清除,但是ED1+的细胞却只能被部分清除,清除的比率取决于前体/成熟巨噬细胞的比例。

Q: Clodronate liposomes没有达到期望的效果?

A:氯磷酸脂质体都是大批量的生产,每批次都会对氯磷酸的浓度以及一些可能的污染物进行检测,确保无误后才会销售给客户。另外,脂质体容易收到温度的影响。正确的运输和储存方式是4~8℃,悬液既不能被冻存,也不能被加热超过30℃。自收到货物起,需要在3个月内使用完,不然有可能会影响使用的效果。

Q:在静脉注射时,能不能增加注射的体积?

A:静脉注射时需要依据动物的体重,不能超过0.1ml/10mg,但是进行腹腔注射可以适当的增加注射的体积。皮下注射需要根据注射位点的容量进行决定。

货号 品名 包装 目录价 * 品牌
78cl005ml Clodronate Liposomes 5ml 4000  2400  clodronaiposomes
78cl010ml Clodronate Liposomes 10ml 7500  4500  clodronaiposomes
78cl015ml Clodronate Liposomes 15ml 9955  5973  clodronaiposomes
78cl020ml Clodronate Liposomes 20ml 11842  7105  clodronaiposomes
78cl025ml Clodronate Liposomes 25ml 13557  8134  clodronaiposomes
78cl030ml Clodronate Liposomes 30ml 15272  9163  clodronaiposomes
78cl040ml Clodronate Liposomes 40ml 18702  11221  clodronaiposomes
78cl050ml Clodronate Liposomes 50ml 22132  13279  clodronaiposomes
78cl060ml Clodronate Liposomes 60ml 24704  14823  clodronaiposomes
78cl070ml Clodronate Liposomes 70ml 27277  16366  clodronaiposomes
78cl080ml Clodronate Liposomes 80ml 29849  17910  clodronaiposomes
78cl090ml Clodronate Liposomes 90ml 31564  18939  clodronaiposomes
78cl100ml Clodronate Liposomes 100ml 33279  19968  clodronaiposomes
78clp005ml Clodronate Liposomes and PBS Liposomes 5ml 4000  2400  clodronaiposomes
78clp010ml Clodronate Liposomes and PBS Liposomes 10ml 7500  4500  clodronaiposomes
78clp015ml Clodronate Liposomes and PBS Liposomes 15ml 10830  6498  clodronaiposomes
78clp020ml Clodronate Liposomes and PBS Liposomes 20ml 14000  8400  clodronaiposomes
78clp025ml Clodronate Liposomes and PBS Liposomes 25ml 17500  10500  clodronaiposomes
78clp030ml Clodronate Liposomes and PBS Liposomes 30ml 20245  12147  clodronaiposomes
78clp040ml Clodronate Liposomes and PBS Liposomes 40ml 24704  14823  clodronaiposomes
78clp050ml Clodronate Liposomes and PBS Liposomes 50ml 28992  17395  clodronaiposomes
78clp060ml Clodronate Liposomes and PBS Liposomes 60ml 32422  19453  clodronaiposomes
78clp070ml Clodronate Liposomes and PBS Liposomes 70ml 35852  21511  clodronaiposomes
78clp080ml Clodronate Liposomes and PBS Liposomes 80ml 38424  23055  clodronaiposomes
78clp090ml Clodronate Liposomes and PBS Liposomes 90ml 40997  24598  clodronaiposomes
78clp100ml Clodronate Liposomes and PBS Liposomes 100ml 43569  26142  clodronaiposomes

 

Hieff Trans<sup>® </sup>悬浮细胞专用脂质体核酸转染试剂 脂质体转染试剂

Hieff Trans<sup>® </sup>悬浮细胞专用脂质体核酸转染试剂 脂质体转染试剂

产品说明书

FAQ

COA

已发表文献

产品描述
Hieff Trans® 悬浮细胞专用脂质体核酸转染试剂是一种阳离子脂质体转染试剂,经优化专门用于悬浮细胞的转染,且适用于DNARNA和寡核苷酸的转染,对大多数真核细胞具有很高的转染效率。
Hieff Trans® 悬浮细胞专用脂质体核酸转染试剂以无菌的液体形式提供。

 

运输与保存方法

冰袋(wet ice)运输。产品2-8ºC保存,一年有效。不可冷冻!

 

注意事项
1. 为得到最优的转染效率,请先用无血清培养基(如OPTI-MEM I培养基)稀释Hieff Trans® Suspension Cell-Free Liposomal Transfection Reagent(以下简称Hieff Trans®),之后与DNARNA做混合。
2. 使用高质量的DNARNA有助于获得较高的转染效率,务必确保质粒的高纯度和无菌状态,彻底去除质粒提取过程中可能残留的苯酚和高盐,因为上述酚类会对细胞造成损失,而高盐分会干扰Hieff Trans®复合物的形成。质粒中的内毒素也是转染的大敌,务必去除。

3. 转染时培养基中不能添加抗生素。

4. 阳离子脂质体应该在2-8保存,要注意避免多次反复长时间开盖,因为可能会导致脂质体氧化而影响转染效率。

5. 需优化DNA浓度和阳离子脂质体试剂量以得到最大的转染效率。DNAHieff Trans®的比例,通常推荐是1:2 1:3

推荐转染条件(以293悬浮培养细胞,质粒DNA转染为例)

最终转染体积:30 mL

转染细胞数目:3×107 cell(最终细胞密度:1×106  cell/mL),转染之前需要确保细胞健康,细胞活力>90%

质粒DNA量:20–40 μg (typically use 30 μg)

转染试剂量:40–80 μL (typically use 60 μL). Use 2 μL Hieff Trans® per 1 μg of plasmid DNA transfected.

操作步骤293悬浮细胞)

使用以下步骤在30 mL总体系内转染293细胞,转染过程中生长培养基内不要添加抗生素,否则会降低转染效率。转染过程请同时设置阳性对照组和阴性对照组(无DNA,无Hieff Trans®)。

【注】对于其他的培养总体系,按比例放大或者缩小各组分用量。
1. 转染当天,在配制复合物之前,准备转染需要的细胞量【见上推荐转染条件,即28 mL 生长培养基内加入3×107 cell,台盼蓝排斥法确定细胞活力和小量细胞成团量。剧烈漩涡混匀45 s以打破细胞团,并用计数器测定总细胞量。细胞活力需>90%。】【注】:为了达到最优效率,确保单细胞悬液。

2. 按照以下方法配制Hieff Trans®-DNA复合物,
a, 用无血清培养基(如OPTI-MEM I)稀释30 μg质粒DNA,使其终体积为1 mL
b, 用无血清培养基(如OPTI-MEM I)稀释60 μL Hieff Trans®,使其终体积为1 mL;轻轻混匀,于室温孵育5 min;【注意】:过长时间孵育会降低效率。
c, 孵育5 min之后,将稀释的质粒DNA加入稀释的Hieff Trans®,使其总体积为2 mL。轻轻混匀。
d, 室温孵育20-30 min,使得DNA-Hieff Trans®复合物形成。此时溶液可能会混浊,但不会影响转染。
【注意】DNA-脂质体复合物室温至少稳定保存5 h
3. 孵育完全后,将2 mL DNA-Hieff Trans®复合物加入28mL 293悬浮细胞的生长培养基内,使得细胞最终的密度约为1×106 cell/mL。对于阴性对照,用2 mL无血清培养基(如OPTI-MEM I)替换。
4. 37℃,5% CO2轨道摇床培养,转速125 rpm,直至进行转基因表达分析,无需去掉复合物或更换培养基。然而,可能有必要在4-6 h后更换生长培养基,不会降低转染活性。

HB230503

 

Q:我们转染试剂转染后需要换液吗?

A:对于换液可以区分两种情况;1、转染之前如果没有换液应在转染 6 小时左右后换液,以保证细胞生长所需营养,2、如果转染之前如果有换液,可以按照平时等到培养基出现营养不足时换液。

Q:转染试剂转单个质粒和多质粒共转的效率如何?

A:单转效率对于验证过的细胞效率都是很好,可以参考 FAQ-验证过的细胞系,对于共转由于要涉及到质粒的混合比例和质粒与转染试剂的添加比例问题,因此具体的效率需要客户做相应的验证方可。

Q:转染试剂可以冻存吗?

A:不可以冻存,因为转染试剂是一种脂质体阳离子转染试剂,由于脂质体是不能在低温下冻存, 因此转染试剂最好是 4 度储存,保持最好的转染效能。

[1] Cheng Y, Liu Y, Shi S, et al. Functional Characterization of Duck STING in IFN-β Induction and Anti-H9N2 Avian Influenza Viruses Infections. Front Immunol. 2019;10:2224. Published 2019 Sep 18. doi:10.3389/fimmu.2019.02224(IF:4.716)
[2] Li J, Peng S, Zhong L, et al. Identification and validation of a regulatory mutation upstream of the BMP2 gene associated with carcass length in pigs. Genet Sel Evol. 2021;53(1):94. Published 2021 Dec 14. doi:10.1186/s12711-021-00689-0(IF:4.297)

产品描述
Hieff Trans® 悬浮细胞专用脂质体核酸转染试剂是一种阳离子脂质体转染试剂,经优化专门用于悬浮细胞的转染,且适用于DNARNA和寡核苷酸的转染,对大多数真核细胞具有很高的转染效率。
Hieff Trans® 悬浮细胞专用脂质体核酸转染试剂以无菌的液体形式提供。

 

运输与保存方法

冰袋(wet ice)运输。产品2-8ºC保存,一年有效。不可冷冻!

 

注意事项
1. 为得到最优的转染效率,请先用无血清培养基(如OPTI-MEM I培养基)稀释Hieff Trans® Suspension Cell-Free Liposomal Transfection Reagent(以下简称Hieff Trans®),之后与DNARNA做混合。
2. 使用高质量的DNARNA有助于获得较高的转染效率,务必确保质粒的高纯度和无菌状态,彻底去除质粒提取过程中可能残留的苯酚和高盐,因为上述酚类会对细胞造成损失,而高盐分会干扰Hieff Trans®复合物的形成。质粒中的内毒素也是转染的大敌,务必去除。

3. 转染时培养基中不能添加抗生素。

4. 阳离子脂质体应该在2-8保存,要注意避免多次反复长时间开盖,因为可能会导致脂质体氧化而影响转染效率。

5. 需优化DNA浓度和阳离子脂质体试剂量以得到最大的转染效率。DNAHieff Trans®的比例,通常推荐是1:2 1:3

推荐转染条件(以293悬浮培养细胞,质粒DNA转染为例)

最终转染体积:30 mL

转染细胞数目:3×107 cell(最终细胞密度:1×106  cell/mL),转染之前需要确保细胞健康,细胞活力>90%

质粒DNA量:20–40 μg (typically use 30 μg)

转染试剂量:40–80 μL (typically use 60 μL). Use 2 μL Hieff Trans® per 1 μg of plasmid DNA transfected.

操作步骤293悬浮细胞)

使用以下步骤在30 mL总体系内转染293细胞,转染过程中生长培养基内不要添加抗生素,否则会降低转染效率。转染过程请同时设置阳性对照组和阴性对照组(无DNA,无Hieff Trans®)。

【注】对于其他的培养总体系,按比例放大或者缩小各组分用量。
1. 转染当天,在配制复合物之前,准备转染需要的细胞量【见上推荐转染条件,即28 mL 生长培养基内加入3×107 cell,台盼蓝排斥法确定细胞活力和小量细胞成团量。剧烈漩涡混匀45 s以打破细胞团,并用计数器测定总细胞量。细胞活力需>90%。】【注】:为了达到最优效率,确保单细胞悬液。

2. 按照以下方法配制Hieff Trans®-DNA复合物,
a, 用无血清培养基(如OPTI-MEM I)稀释30 μg质粒DNA,使其终体积为1 mL
b, 用无血清培养基(如OPTI-MEM I)稀释60 μL Hieff Trans®,使其终体积为1 mL;轻轻混匀,于室温孵育5 min;【注意】:过长时间孵育会降低效率。
c, 孵育5 min之后,将稀释的质粒DNA加入稀释的Hieff Trans®,使其总体积为2 mL。轻轻混匀。
d, 室温孵育20-30 min,使得DNA-Hieff Trans®复合物形成。此时溶液可能会混浊,但不会影响转染。
【注意】DNA-脂质体复合物室温至少稳定保存5 h
3. 孵育完全后,将2 mL DNA-Hieff Trans®复合物加入28mL 293悬浮细胞的生长培养基内,使得细胞最终的密度约为1×106 cell/mL。对于阴性对照,用2 mL无血清培养基(如OPTI-MEM I)替换。
4. 37℃,5% CO2轨道摇床培养,转速125 rpm,直至进行转基因表达分析,无需去掉复合物或更换培养基。然而,可能有必要在4-6 h后更换生长培养基,不会降低转染活性。

HB230503

 

Q:我们转染试剂转染后需要换液吗?

A:对于换液可以区分两种情况;1、转染之前如果没有换液应在转染 6 小时左右后换液,以保证细胞生长所需营养,2、如果转染之前如果有换液,可以按照平时等到培养基出现营养不足时换液。

Q:转染试剂转单个质粒和多质粒共转的效率如何?

A:单转效率对于验证过的细胞效率都是很好,可以参考 FAQ-验证过的细胞系,对于共转由于要涉及到质粒的混合比例和质粒与转染试剂的添加比例问题,因此具体的效率需要客户做相应的验证方可。

Q:转染试剂可以冻存吗?

A:不可以冻存,因为转染试剂是一种脂质体阳离子转染试剂,由于脂质体是不能在低温下冻存, 因此转染试剂最好是 4 度储存,保持最好的转染效能。

[1] Cheng Y, Liu Y, Shi S, et al. Functional Characterization of Duck STING in IFN-β Induction and Anti-H9N2 Avian Influenza Viruses Infections. Front Immunol. 2019;10:2224. Published 2019 Sep 18. doi:10.3389/fimmu.2019.02224(IF:4.716)
[2] Li J, Peng S, Zhong L, et al. Identification and validation of a regulatory mutation upstream of the BMP2 gene associated with carcass length in pigs. Genet Sel Evol. 2021;53(1):94. Published 2021 Dec 14. doi:10.1186/s12711-021-00689-0(IF:4.297)