Protein isolation from mucus

[GeneMan]

Hi all

 

Anyone out there know how to isolate total protein from respiratory mucus, namely sinus snot? I have a long method that removes cells (dint want them in there, just mucus protein!), using DTT, followed by an acetone precipitation. Its way labor intensive and uses allot of acetone to work. I want a kit or a nice short method that takes into accout removing cells. Anything???

 

Cheers

GeneMan

[fiveworlds]

1. add to beaker

2, add water

3. add washing up liquid(type matters depending on which you use more of the cell is dissolved into solution pick carefully)

4. stir

5. wait

6. centrifuge

7. chromatography/gel electrophoresis

 

 

There is also some total protein webpages http://labtestsonline.org/understanding/analytes/tp/tab/test/ that talk about protein electrophoresis. A doctor should be able to distinguish between the proteins.

Edited July 9, 2014 by fiveworlds

[CharonY]

1. add to beaker

2, add water

3. add washing up liquid(type matters depending on which you use more of the cell is dissolved into solution pick carefully)

4. stir

5. wait

6. centrifuge

7. chromatography/gel electrophoresis

 

 

There is also some total protein webpages http://labtestsonline.org/understanding/analytes/tp/tab/test/ that talk about protein electrophoresis. A doctor should be able to distinguish between the proteins.

 

That is not going to work well.

 

For OP, acetone precipitation is pretty much one of the fastest ways to concentrate and (to some extent) desalt proteins. With regards to cell removal it is not clear to me how lengthy your procedure is but typically involves centrifugation and/or filtration. In some cases there is even further extraction to remove polysaccharides but how crude the extraction could be really depends on what type of analyses you want to do downstream.

[fiveworlds]

 

That is not going to work well.

 

For OP, acetone precipitation is pretty much one of the fastest ways to concentrate and (to some extent) desalt proteins. With regards to cell removal it is not clear to me how lengthy your procedure is but typically involves centrifugation and/or filtration. In some cases there is even further extraction to remove polysaccharides but how crude the extraction could be really depends on what type of analyses you want to do downstream.

It would be the only method I know. If you know of a better method i would be more than willing to learn it. There is also http://en.wikipedia.org/wiki/Serum_protein_electrophoresis but that involves using blood not mucus

Edited July 9, 2014 by fiveworlds

[GeneMan]

Thanks Charon Y and FiveWorlds

 

Im good for electrophoresis, thanks Charon Y

 

I think FiveWorlds is on to some thing here. My method actually works very well with acetone, its just huge due to the volumes needed after atting the DTT solution to adequately remove the cells (happy to supply FiveWorld, although is in the lab boom atm so will need to transpose to doc.) Hence when you add the acetone for the correct volume to voluem ratio, it gets volumous and and difficult to manage a range of samples/patients. Trying to process 10's of snots.

 

Ideally I want a kit, eg there are saliva kits, coz 1. we can afford it, and 2. my time is costly (although I suspect the kit is worth more than my time (sigh)).

 

Chees for the advice though people.

GeneMan