g*********5
发帖数: 2533 | 1 I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
100-120 kDa on trisglycine gels...
and I know this protein is phosphyralted protein, but different is so big.
and santa cruz antibody give a lot band with western, and even that a lot
ppls use santa cluz antibody do immunofluoresence...
and some labs use their homemade protein purified antibody and go 68kDa band
... and some ppl said Nrf2 at 53kDa on their paper...
confuse me.... |
b*****n
发帖数: 1841 | 2 窥一斑知豹，这个是当今科研混乱的一个缩影。
is
band
【在 g*********5 的大作中提到】
: I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
: 100-120 kDa on trisglycine gels...
: and I know this protein is phosphyralted protein, but different is so big.
: and santa cruz antibody give a lot band with western, and even that a lot
: ppls use santa cluz antibody do immunofluoresence...
: and some labs use their homemade protein purified antibody and go 68kDa band
: ... and some ppl said Nrf2 at 53kDa on their paper...
: confuse me....
|
a*****x
发帖数: 901 | 3 Did you do the control of over-expression and knock-down? If not, you can't
never tell whether the band you saw represents your protein.
is
band
【在 g*********5 的大作中提到】
: I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
: 100-120 kDa on trisglycine gels...
: and I know this protein is phosphyralted protein, but different is so big.
: and santa cruz antibody give a lot band with western, and even that a lot
: ppls use santa cluz antibody do immunofluoresence...
: and some labs use their homemade protein purified antibody and go 68kDa band
: ... and some ppl said Nrf2 at 53kDa on their paper...
: confuse me....
|
u**********d
发帖数: 573 | 4 过表达用标签看看全长蛋白的位置到底在哪里？
也许修饰也许splicing导致条带位置在不同状态下变化。
is
band
【在 g*********5 的大作中提到】
: I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
: 100-120 kDa on trisglycine gels...
: and I know this protein is phosphyralted protein, but different is so big.
: and santa cruz antibody give a lot band with western, and even that a lot
: ppls use santa cluz antibody do immunofluoresence...
: and some labs use their homemade protein purified antibody and go 68kDa band
: ... and some ppl said Nrf2 at 53kDa on their paper...
: confuse me....
|
q*******8
发帖数: 768 | 5 这种事也是常有的，有时候也是解释不清。PHOSPHORYLATION SHIFT有时候也是很明显
的。但也有可能是WESTERN假阳性。你呐，要不割下来去测个序。有时间的话呢，给蛋
白加个TAG，然后用TAG纯化下来再做WESTERN |
q*******8
发帖数: 768 | 6 不能这么说，不同实验室做出来不同结果是很正常的，也是可以解释的，比如之前某人
做的时候用了一种要抑制剂以为是特定的，后来发现抑制剂有别的功能；有的人药品用
酒精溶，有的人用DMSO溶，有时候会有截然不同的结果。。。这是科研向前走的必由之
路呀~
【在 b*****n 的大作中提到】
: 窥一斑知豹，这个是当今科研混乱的一个缩影。
:
: is
: band
|
i***0
发帖数: 160 | 7 哥们发贴之前能不能先读一遍，好歹改一下，咱好歹也是学术版啊。不然就发中文吧。 |
D***a
发帖数: 516 | 8 有时候我也搞不懂，同样一个蛋白，不同公司的抗体说明书里给的size就不一样。
is
band
【在 g*********5 的大作中提到】
: I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
: 100-120 kDa on trisglycine gels...
: and I know this protein is phosphyralted protein, but different is so big.
: and santa cruz antibody give a lot band with western, and even that a lot
: ppls use santa cluz antibody do immunofluoresence...
: and some labs use their homemade protein purified antibody and go 68kDa band
: ... and some ppl said Nrf2 at 53kDa on their paper...
: confuse me....
|
f*****f
发帖数: 195 | 9 这个很正常，最常见的情况是至少有个公司是错的。在极少且特殊的情况，蛋白有不同
的剪切或修饰如sumo化，不同抗体识别不一样结构。
【在 D***a 的大作中提到】
: 有时候我也搞不懂，同样一个蛋白，不同公司的抗体说明书里给的size就不一样。
:
: is
: band
|
S*********s
发帖数: 304 | 10 如果你做一下recombinant protein, 你就知道size了
目前的很多用santa cruz抗体做的IF和WB都是不可信的。
通过overexpression 和siRNA knockdown, 只有两条band是真的。
abcam，santa cruz,cell signaling卖的抗体里能用的似乎只有一个是能用的。
is
band
【在 g*********5 的大作中提到】
: I am studing a protein Nrf2 now, and predict size should be 68kD, but it is
: 100-120 kDa on trisglycine gels...
: and I know this protein is phosphyralted protein, but different is so big.
: and santa cruz antibody give a lot band with western, and even that a lot
: ppls use santa cluz antibody do immunofluoresence...
: and some labs use their homemade protein purified antibody and go 68kDa band
: ... and some ppl said Nrf2 at 53kDa on their paper...
: confuse me....
|
