SDS-PAGE showing 1µg purified Ephrin-B2 (EFNB2) protein. The molecular mass is approximately 35-40 kDa due to glycosylation.

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HUMAN EPHRIN-B2 (EFNB2), HIS-TAG
Recombinant EFNB2 protein produced in HEK293 cells and purified from culture supernatant. Protein contains an C-terminal 6x His-tag.
PRODUCT DETAILS – HUMAN EPHRIN-B2 (EFNB2), HIS-TAG
- Recombinant EFNB2 protein produced from HEK293 cells (NCBI Accession Number: NP_004084.1).
- Includes amino acids 1-226 and a C-terminal His-tag.
- Greater than 95% purity by SDS-PAGE and buffered in DPBS, pH7.4.
BACKGROUND
Human Ephrin-B2 is encoded by the EFNB2 gene, a EFNB class ephrin which binds to the EPHB4 and EPHA3 receptors (Bonaldo, et al., 1994). EFNB2 is a member of the family of transmembrane-anchored ligands of the ephrin receptor tyrosine kinase family, the largest subgroup of receptor tyrosine kinases known (Poliakov, et al., 2004; Pasquale, 2004) and are type I membrane glycoproteins.
Ephrin receptors and their ligands are known to participate in cell-cell interactions, including those of vascular endothelial cells, and are modulators of cell migration in remodeling events, especially in the nervous system. EFNB2 expression is seen in most human tissues to varying degrees.
Hendra virus (HeV) and Nipah virus (NiV) belong to the genus Henipavirus of the family Paramyxoviridae and are unique in that they exhibit a broad species tropism and cause fatal disease in both animals and humans. They infect cells through the same pH-independent membrane fusion process mediated by their fusion and attachment glycoproteins.
EFNB2 has been shown to serve as a functional receptor for both Hendra virus (HeV) and Nipah virus (NiV). Non-permissive cells transfected with a vector encoding EFNB2 became permissive for HeV- and NiV-mediated cell fusion as well as infection by live virus. Additionally, recombinant EFNB2 could potently block fusion and infection and bind recombinant HeV and NiV attachment glycoproteins with high affinity (Bonaparte, et al., 2005).
REFERENCES
- Bonaldo, M. F. et al., 1994. Selection of cDNAs using chromosome-specific genomic clones: application to human chromosome 13. Human Molecular Genetics, 3(9), p. 1663–73.
- Bonaparte, M. I. et al., 2005. Ephrin-B2 ligand is a functional receptor for Hendra virus and Nipah virus. Proc Natl Acad Sci U S A., 102(30), pp. 10652-7.
- Pasquale, E. B., 2004. Eph-ephrin promiscuity is now crystal clear. Nat Neurosci., 7(5), pp. 417-8.
- Poliakov, a., Cotrina, M. & Wilkinson, D. G., 2004. Diverse roles of eph receptors and ephrins in the regulation of cell migration and tissue assembly. Dev Cell, 7(4), pp. 465-80.
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将来可能会出现基于免疫方法的试剂盒,不过因为H7N9是个新病毒,爆发至今不足一月,这么快时间还来不及生产抗体,所以目前的检测试剂盒只能是PCR检测试剂盒。
查到了:
4月7日,上海之江生物科技有限公司官方微博称,该公司成功研制禽流感H7N9(2013)核酸测定试剂盒(荧光PCR法),是针对国内此次H7N9病毒最早研制成功的产品,也是国内目前唯一供应的成品化试剂盒。
它一般在医院、制药企业使用。
试剂盒使用示例: 试剂盒的产生正是为了使实验人员能够摆脱繁重的试剂配制及优化过程,所以试剂盒中一般配备有相应的使用说明书
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。
如果用组织DNA提取试剂盒,提出来的就是组织细胞的DNA了