Can anybody help me with protocol for protein extraction from sds polyacrylamide gel for raising hyper immune sera..the gel is used is of 15% strength from virus
0
5 replies to this topic
#2
Adrian K
-
- Global Moderators
-
- 684 posts
Legendary Graduate Beggar
24
Excellent
Excellent
Posted 08 July 2012 - 02:46 PM
Easy and fastest way: you can try electroelution. Can look for commercial kits such as pierce or merck.
There is a protocol on WILEY but I do not have access to it:
http://onlinelibrary...808s46/abstract
Another from SPRINGER (I think is free access)
http://www.springerl...36w023410u816u/
Also, you may consider:
http://onlinelibrary...CO;2-X/abstract
http://www.springerl...&page=4&locus=6
Hope this helps.
There is a protocol on WILEY but I do not have access to it:
http://onlinelibrary...808s46/abstract
Another from SPRINGER (I think is free access)
http://www.springerl...36w023410u816u/
Also, you may consider:
http://onlinelibrary...CO;2-X/abstract
http://www.springerl...&page=4&locus=6
Hope this helps.
Expecting the world to treat you fairly because you are a good person is like expecting the lion not to attack you because you are a vegetarian.
..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
#3
mdfenko
-
- Active Members
-
- 2,251 posts
an elder
79
Excellent
Excellent
Posted 09 July 2012 - 08:15 AM
you can excise the band, mash it and inject it (that's the way we did it). there is no need to remove the gel for inoculation.
talent does what it can
genius does what it must
i do what i get paid to do
genius does what it must
i do what i get paid to do
#4
biozweistein22
-
- Active Members
-
- 8 posts
member
0
Neutral
Neutral
Posted 09 July 2012 - 12:36 PM
First of all, mdfenko is right. The easiest way is to excise the band itself and mash it in PBS, but this will raise a huge immune response and may kill your animal because the gel is toxic, since you'll have to stain and destain it to see your band. If you do that, use coomassie and destain with 2-propanol instead of methanol as it is milder.
#5
mdfenko
-
- Active Members
-
- 2,251 posts
an elder
79
Excellent
Excellent
Posted 11 July 2012 - 05:43 AM
biozweistein22, on 09 July 2012 - 12:36 PM, said:
First of all, mdfenko is right. The easiest way is to excise the band itself and mash it in PBS, but this will raise a huge immune response and may kill your animal because the gel is toxic, since you'll have to stain and destain it to see your band. If you do that, use coomassie and destain with 2-propanol instead of methanol as it is milder.
talent does what it can
genius does what it must
i do what i get paid to do
genius does what it must
i do what i get paid to do
#6
biozweistein22
-
- Active Members
-
- 8 posts
member
0
Neutral
Neutral
Posted 11 July 2012 - 06:44 AM
Both coomassie and destain contain methanol, at least in our lab, that's why I said that the gel's staining and destaining makes it toxic. To avoid that, we wash the gel as you said using PBS before mashing it, and also use 2-propanol instead of methanol to make the stain-destain. In my experience it is much safer that way. Hope it works....
