J*n
2 楼
I have two proteins, and the binding is not very strong. IP and pull-down
assay showed positive results. But we want to have other evidence of the
binding?
Is ITC a good tool to study protein-protein interaction?
And is there any other approaches to study protein-protein interaction? Size
exclusion chromatography? SPR? ....
Many thanks in advance!
assay showed positive results. But we want to have other evidence of the
binding?
Is ITC a good tool to study protein-protein interaction?
And is there any other approaches to study protein-protein interaction? Size
exclusion chromatography? SPR? ....
Many thanks in advance!
p*n
3 楼
spr
itc is not good for weak interactions.
sec may work. get a hplc sec column.
Size
【在 J*****n 的大作中提到】
: I have two proteins, and the binding is not very strong. IP and pull-down
: assay showed positive results. But we want to have other evidence of the
: binding?
: Is ITC a good tool to study protein-protein interaction?
: And is there any other approaches to study protein-protein interaction? Size
: exclusion chromatography? SPR? ....
: Many thanks in advance!
itc is not good for weak interactions.
sec may work. get a hplc sec column.
Size
【在 J*****n 的大作中提到】
: I have two proteins, and the binding is not very strong. IP and pull-down
: assay showed positive results. But we want to have other evidence of the
: binding?
: Is ITC a good tool to study protein-protein interaction?
: And is there any other approaches to study protein-protein interaction? Size
: exclusion chromatography? SPR? ....
: Many thanks in advance!
e*s
4 楼
Sec won"t work for weak association.
Weak protein-protein interaction is always hard to confirm.
If one protein is small, you may try FP.
【在 p*****n 的大作中提到】
: spr
: itc is not good for weak interactions.
: sec may work. get a hplc sec column.
:
: Size
Weak protein-protein interaction is always hard to confirm.
If one protein is small, you may try FP.
【在 p*****n 的大作中提到】
: spr
: itc is not good for weak interactions.
: sec may work. get a hplc sec column.
:
: Size
l*1
5 楼
FRET
k*0
6 楼
Your answer is quite right.
【在 e****s 的大作中提到】
: Sec won"t work for weak association.
: Weak protein-protein interaction is always hard to confirm.
: If one protein is small, you may try FP.
【在 e****s 的大作中提到】
: Sec won"t work for weak association.
: Weak protein-protein interaction is always hard to confirm.
: If one protein is small, you may try FP.
A*w
7 楼
For ITC, you need milligrams of protein.
SPR, you need much less.
Another easy one is chemical cross-linking...
SPR, you need much less.
Another easy one is chemical cross-linking...
H*g
8 楼
There are many ways to show protein-protein interaction; However, I would
suggest u to forget any additional in vitro assays. You already got IP and
pull-down, so do not waste time on other in vitro ones.
In your case, if I were the reviewer, I would directly ask for evidence from
other perspectives, in addition to all those in vitro assays. I would ask
for assays testing in vivo, real-time interactions, such as BiFC or FRET, as
well as any evidence demonstrating genetic interaction to support that the
physical interaction is biological meaningful.
Size
【在 J*****n 的大作中提到】
: I have two proteins, and the binding is not very strong. IP and pull-down
: assay showed positive results. But we want to have other evidence of the
: binding?
: Is ITC a good tool to study protein-protein interaction?
: And is there any other approaches to study protein-protein interaction? Size
: exclusion chromatography? SPR? ....
: Many thanks in advance!
suggest u to forget any additional in vitro assays. You already got IP and
pull-down, so do not waste time on other in vitro ones.
In your case, if I were the reviewer, I would directly ask for evidence from
other perspectives, in addition to all those in vitro assays. I would ask
for assays testing in vivo, real-time interactions, such as BiFC or FRET, as
well as any evidence demonstrating genetic interaction to support that the
physical interaction is biological meaningful.
Size
【在 J*****n 的大作中提到】
: I have two proteins, and the binding is not very strong. IP and pull-down
: assay showed positive results. But we want to have other evidence of the
: binding?
: Is ITC a good tool to study protein-protein interaction?
: And is there any other approaches to study protein-protein interaction? Size
: exclusion chromatography? SPR? ....
: Many thanks in advance!
J*n
9 楼
Thank you all for the kindly replies.
Two of my proteins, one is 44 kd and another is 67, therefore I do not think
it is appropriate to use FP.
ITC did not give me positive results.
SEC showed binding but the peaks are ambiguous.
We might try MALS...not sure if it will be successful.
Actually, to be more specific, we want to check the binding stoichiometry of
these two proteins. Any suggestion?
Many thanks!
Two of my proteins, one is 44 kd and another is 67, therefore I do not think
it is appropriate to use FP.
ITC did not give me positive results.
SEC showed binding but the peaks are ambiguous.
We might try MALS...not sure if it will be successful.
Actually, to be more specific, we want to check the binding stoichiometry of
these two proteins. Any suggestion?
Many thanks!
相关阅读
那位解释一下，这篇文章为什么能发SCIENCE有发过Mol Microbiol的吗？某快顶不住了PGP9.5和neuralfilament 200抗体识别神经轴突有何差别？国内干临床的医生拿医学学士来美国做博士后，这也太爽啦谢晓亮小组报告新技术有望大幅增加试管婴儿成功率是重新申请CS还是一边读PhD，一边辅修个学位？Kevin White 到底有多牛？我想发布个生信软件，不知要在发表文章之前还是之后？做lentivirus防护不到位的话，被感染了有症状吗求助打听博后老板：lewis Cantley/Tyle Jacks/Kang yibin/3PUniversity of Florida农学系的offer怎么还没下来啊！真的要跳入火坑吗？请大家介绍生物论坛网站要么早归，要么别归，不要温水煮青蛙四年，换了5个地方（转载）有没有做mouse intestine的，帮我看一下这个图片。申请NIH GRANT 对TITLE有什么要求吗阶段性 小解脱帮人求文献，比较老的求教Lentivirus shRNA knock-down实验