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how to avoid the keratin contamination form 2-DE gel - kindly help me!!!!!! (Jun/05/2008 )

Hi everyone,

i have run 2-DE gel with extra precaution to avoid keratin contamination but as i sent the spot for MALDI, then technician has told me that there is enormous keratin contamination. and now i am in dillemma. as i have left no stone unturned to avoid keratin, even though the keratin are present.


kindly suggest me something regarding the key strategies of keratin?

Can it be possible that i cut some other spot and send it for MALDI?

is there any method to remove the keratin form the gel? or do i need to again run the gel?


your kind help would be highly appreciated.


thanx in advance.

-pBS-

QUOTE (pBS @ Jun 5 2008, 09:32 PM)
Hi everyone,

i have run 2-DE gel with extra precaution to avoid keratin contamination but as i sent the spot for MALDI, then technician has told me that there is enormous keratin contamination. and now i am in dillemma. as i have left no stone unturned to avoid keratin, even though the keratin are present.


kindly suggest me something regarding the key strategies of keratin?

Can it be possible that i cut some other spot and send it for MALDI?

is there any method to remove the keratin form the gel? or do i need to again run the gel?


your kind help would be highly appreciated.


thanx in advance.


external keratins are from epidermal skin and hair; prepare ALL solutions, material and chambers for 2D under excellent clean conditions; try to simulate clean room conditions

-The Bearer-

QUOTE (The Bearer @ Jun 6 2008, 01:31 AM)
QUOTE (pBS @ Jun 5 2008, 09:32 PM)
Hi everyone,

i have run 2-DE gel with extra precaution to avoid keratin contamination but as i sent the spot for MALDI, then technician has told me that there is enormous keratin contamination. and now i am in dillemma. as i have left no stone unturned to avoid keratin, even though the keratin are present.


kindly suggest me something regarding the key strategies of keratin?

Can it be possible that i cut some other spot and send it for MALDI?

is there any method to remove the keratin form the gel? or do i need to again run the gel?


your kind help would be highly appreciated.


thanx in advance.


external keratins are from epidermal skin and hair; prepare ALL solutions, material and chambers for 2D under excellent clean conditions; try to simulate clean room conditions




thanx for kind reply.

would you please tell me how to simulate the room condition. Is your meaning to sterilize the room with disinfectant?

I use glassware after washing them with detergenta and rinsing them a number of times with miiliQ water. Is there any treatment for removing the keratin form the glassware?

looking for response.

-pBS-

There are many links here which describe the problem and solutions:
http://search.vadlo.com/b/q?sn=158621799&a...ation&rel=0
hth/

-cellcounter-

the way we avoid keratins in our gels and westerns is to omit the reducing agent from the sample buffer.

this can't be done for all proteins but will work for some.

-mdfenko-