SDS-PAGE showing 1µg purified DC-SIGN protein (indicated by arrow).

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HUMAN DC-SIGN, HIS-TAG
Recombinant human dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) with N-terminal His-tag.
PRODUCT DETAILS – HUMAN DC-SIGN, HIS-TAG
- Recombinant human dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) expressed from HEK293 cells (NCBI Accession Number: NP_066978.1).
- Includes amino acids 59-404 and an N-terminal His-tag.
- Greater than 95% purity by SDS-PAGE and buffered in DPBS, pH7.4.
BACKGROUND
Human DC-SIGN (Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin) also known as CD209 (Cluster of Differentiation 209) is encoded by the CD209 gene (Curtis, et al., 1992). It is a 44-kDa type II integral membrane protein. The protein is organized into three distinct domains: an N-terminal transmembrane domain, a tandem-repeat neck domain and C-type lectin carbohydrate recognition domain. The extracellular region consisting of the C-type lectin and neck domains has a dual function as a pathogen recognition receptor and a cell adhesion receptor by binding carbohydrate ligands on the surface of microbes and endogenous cells. The neck region is important for homo-oligomerization which allows the receptor to bind multivalent ligands with high avidity. Variations in the number of 23 amino acid repeats in the neck domain of this protein are rare but have a significant impact on ligand binding ability.
DC-SIGN is present on the surface of macrophages (and dendritic cells) and recognises and binds to mannose type carbohydrates (high-mannose-containing envelope glycoproteins) which are commonly found on viruses, bacteria and fungi. This binding interaction then activates phagocytosis (McGreal, et al., 2005).
DC-SIGN is a receptor for several viruses including HIV, Hepatitis C, Dengue, Ebola and CMV (Lozach, et al., 2004; Lozach, et al., 2007; Geijtenbeek, et al., 2000) and binding allows them to infect T-cells from dendritic cells. This is an essential process for HIV infection (Wu, et al., 2002) where in the initial stages of dendritic cell infection, the HIV gp120 protein causes co-internalization of DC-SIGN and HIV virions. The dendritic cell then migrates to the cognate lymphoid organ, whereupon recycling of the DC-SIGN/HIV virion complex to the cell periphery facilitates HIV infection of CD4+ T cells by interaction between DC-SIGN and ICAM-3 (van den Berg & Geijtenbeek, 2013).
Dengue virus (serotypes 1-4) also use DC-SIGN to infect dendritic cells.THP-1 cells have been shown to become susceptible to Dengue infection after transfection of DC-SIGN (or its homologue L-SIGN). However, infection of dendritic cells is blocked by anti–DC-SIGN antibodies, but not by antibodies to other molecules on these cells. Viruses produced by dendritic cells are subsequently infectious for DC-SIGN (and L-SIGN) bearing THP-1 cells and other permissive cell lines (Tassaneetrithep, et al., 2003).
DC-SIGN (and L-SIGN) also bind Ebola virus glycoproteins and dendritic cells expressing DC-SIGN are more efficiently infected by Ebola (Alvarez, et al., 2002). CMV envelope glycoprotein B is a viral ligand for DC-SIGN (and L-SIGN). Blocking DC-SIGN with antibodies inhibit dendritic cell infection and conversely expression of DC-SIGN (or L-SIGN) rendering cells permissive to CMV infection (Baribaud, et al., 2002). Therefore, DC-SIGN is a promising target for designing therapies to block pathogen infection.
REFERENCES
- Alvarez, C. P. et al., 2002. C-type lectins mediate cellular entry by Ebola virus in cis and in trans. J Virol., 76(13), pp. 6841-4.
- Baribaud, F. et al., 2002. Human cytomegalovirus binding to DC-SIGN is required for dendritic cell infection and target cell trans-infection. J Virol., 76(18), pp. 9135-42.
- Curtis, B. M., Scharnowske, S. & Watson, A. J., 1992. Sequence and expression of a membrane-associated C-type lectin that exhibits CD4-independent binding of human immunodeficiency virus envelope glycoprotein gp120. Proc. Natl. Acad. Sci. U.S.A., 89(17).
- Geijtenbeek, T. B. et al., 2000. DC-SIGN, a dendritic cell-specific HIV-1-binding protein that enhances trans-infection of T cells. Cell, 100(5), p. 587–97.
- Lozach, P. Y. et al., 2004. C-type lectins L-SIGN and DC-SIGN capture and transmit infectious hepatitis C virus pseudotype particles. J. Biol. Chem., 279(31), p. 32035–45.
- Lozach, P. Y., Burleigh, L., Staropoli, I. & Amara, A., 2007. The C type lectins DC-SIGN and L-SIGN: receptors for viral glycoproteins. Methods Mol. Biol., Volume 379, p. 51–68.
- McGreal, E., Miller, J. & Gordon, S., 2005. Ligand recognition by antigen-presenting cell C-type lectin receptors. Curr Opin Immunol., 17(1), pp. 18-24.
- Tassaneetrithep, B. et al., 2003. DC-SIGN (CD209) Mediates Dengue Virus Infection of Human Dendritic Cells. J Exp Med., 197(7), p. 823–829.
- van den Berg, L. M. & Geijtenbeek, T. B., 2013. Antiviral immune responses by human langerhans cells and dendritic cells in HIV-1 infection. Advances in Experimental Medicine and Biology, Volume 762, p. 45–70.
- Wu, L. et al., 2002. Functional Evaluation of DC-SIGN Monoclonal Antibodies Reveals DC-SIGN Interactions with ICAM-3 Do Not Promote Human Immunodeficiency Virus Type 1 Transmission. J. Virol., 76(12), p. 5905–14.
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如果用组织DNA提取试剂盒,提出来的就是组织细胞的DNA了
短截就是把枝条剪短,主要作用是促使其抽生新梢,增加分枝数目,以保证树势健状和正常结果。短截常用于骨干枝组修剪,结果枝组修剪,和树体局部更新复状。
短截按其长度可分为:
① 中短截:在一年生枝的中部短截,剪后萌发的顶端枝条,长势强,下部枝条长势弱。
② 重短截:剪去一年生枝的2/3。剪后萌发出的枝条较强状,一般用于主侧枝延长头修剪。
③ 重剪:剪去一年生枝的3/4-4/5,剪后萌发出的枝条长势强状,常用于发育枝作延长枝头和徒长果枝,中果枝的修剪。
④极重短截:剪去一年生枝的4/5以上,萌发后的枝条中庸偏状,常用于将发育枝和徒长枝培养结果枝组。
⑤留基部2芽剪:剪后萌发枝条较旺盛,常用于预备枝的修剪。对于幼龄树,树势较旺,以培养良好而牢固的树形结构,提早结果为主要目的,以轻短截,少疏间为主,从始果期到盛果期,主要使桃树多结果,并形成好的树形。
如果PCR产物不是很纯,或者PCR扩增条带比较小,PCR产物前面又有较多引物二聚体时,用胶回收,其余用PCR产物纯化试剂盒。
pcr纯化试剂盒和胶回收试剂盒的区别:
PCR纯化试剂盒:是直接水溶解的PAC产物就可以回收,回收效率高,但是只适合单一条带需要纯化测序的时候使用。
PCR凝胶试剂盒:是在PCR产物是混合物,有多条杂带的情况下,先跑胶将杂带分离,然后在将所要的条带位置的胶切下回收,后者的回收效率低,但是很纯净。
胶回收试剂盒操作步骤:
配制琼脂糖EB凝胶,电泳以分离DNA片段。任何类型或等级的琼脂糖都可以使用。
电泳足够时间后,在紫外灯下小心地把所需的DNA的片段切下来。并尽量去除多余的凝胶。
称取空离心管的重量,切下带目的片段的凝胶装在1.5ml离心管中并称其重量,求出凝胶块的重量,近似地确定其体积。一般情况下,凝胶的密度为1g/ml,于是凝胶的体积与重量的关系可按下面换算:凝胶薄片的重量为0.2g 则其体积为0.2ml;加入等倍凝胶体积的Binding Buffer,把混合物置于55℃~65℃水浴中温浴7min至凝胶完全融化,其间每隔2-3分钟混匀一次;
转移700μl的DNA-琼脂糖溶液到一个HiBindTM DNA柱子,并把柱子装在一个干净的2ml收集管内,室温下,10,000×g离心1min,弃去液体。
将柱子重新套回收集管中,加300μl Binding Buffer至HiBind DNA 柱子中;室温下,10,000×g离心 1分钟,去弃滤出液;这一步相当关键,不要忽略此步。
将柱子重新套回收集管中,加入700μl SPW Wash buffer至HiBind DNA柱子中,室温下,10,000×g离心1分钟,去弃滤出液;注:SPW Wash buffer在使用前必须按瓶子标鉴要求用无水乙醇进行稀释。
将柱子重新套回收集管中,重复加入700μl SPW Wash buffer至HiBind DNA柱子中,室温下,10,000×g离心1分钟,去弃滤出液;
弃去液体,将空柱子重新套回收集管中,10,000×g离心1min以甩干柱基质残余的液体。
这步可以去除柱子基质上残余的乙醇,不要省略此步―――对得到好的DNA产量是十分重要的。
把柱子装在一个干净的1.5ml离心管上,加入30~50μl洗脱液或灭菌水上柱子膜上,10,000×g离心1分钟,离心管中的溶液就是纯化的DNA产物,保存于-20度。
从1~2个T75培养皿的感染细胞培养液中纯化腺病毒
产品简介
本系列试剂盒可快速,高效地从腺病毒感染的细胞培养液中分离纯化腺病毒,相比于传统CsCl超速离心的腺病毒病毒纯化方式(需要24个小时才能完成纯化),本试剂盒可以在1个小时内完成病毒纯化,操作简单,快速,纯化率高,纯化到的病毒颗粒可直接用于下游实验,如细胞和动物感染。
产品特点
?操作简单,快速,可以在1个小时内完成病毒纯化,不依赖于超速离心操作。
?纯化效率高:小量纯化,可从1~2个T75瓶培养的细胞培养液中纯化病毒颗粒高达1x1012VPs
?每个纯化柱,可以重复利用一次,用于纯化相同种类的腺病毒。
保存条件
纯化柱和脱盐柱保存于4℃,其它组分室温保存。
试剂盒组分
Catalog#
V1160-01
Notes
Preps
10
MiniColumns
5
Canbeusedtwice
(Storeat4°C)
Press-OnCap
5
StoreatRT
DesaltingTube*
1
Canberegenarated
(Storeat4°C)
15mLCollectionTube
10
StoreatRT
10xWashBuffer
30mL
StoreatRT
2xElutionBuffer
30mL
StoreatRT
RegenerationBuffer
30mL
StoreatRT
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