Powdered acrylamide versus solution acrylamide - (Oct/01/2008 )
Hi,
I am new to proteomics and I would like to ask whether in your lab you use (1) powdered form acrylamide or (2) Bio-rad or GE liquid form acrylamide solution to cast gel?
It seems that the powdered form acrylamide as well as bis are far too dangerous to use, although it is much cheaper.
And I'd also like to ask whether you wear a face mask or do in a hood or other important precautions (e.g. when adding TEMED) if you cast a gel.
Thx in advance~
I am new to proteomics and I would like to ask whether in your lab you use (1) powdered form acrylamide or (2) Bio-rad or GE liquid form acrylamide solution to cast gel?
It seems that the powdered form acrylamide as well as bis are far too dangerous to use, although it is much cheaper.
And I'd also like to ask whether you wear a face mask or do in a hood or other important precautions (e.g. when adding TEMED) if you cast a gel.
Thx in advance~
Danger and risk are both relative terms. If your lab has the money, I'd go for the pre-mixed reagents. Much simpler, but the trade-off is the extra cost.
I worked in a hood for the powder-based steps, but not for the steps that use liquids. The best advice I was given was to make sure you have everything you need within reach, and keep the workspace clear, so you're less likely to knock things over (this was after I accidently knocked over a bottle of TEMED: it releases a lot of ammonium... what's the emoticon for choking and spluttering?
I am new to proteomics and I would like to ask whether in your lab you use (1) powdered form acrylamide or (2) Bio-rad or GE liquid form acrylamide solution to cast gel?
It seems that the powdered form acrylamide as well as bis are far too dangerous to use, although it is much cheaper.
And I'd also like to ask whether you wear a face mask or do in a hood or other important precautions (e.g. when adding TEMED) if you cast a gel.
Thx in advance~
Acrylamide in any form is a neuro toxin. U can wear mask when u prepare the chemicals and while handling acrylamide in liquid it is advisable to use gloves.
When I worked in an academic lab, we prepared our own 40% stock twice a year so that our exposure to the powder was limited.
Hi, this is a very good idea, can u kindly teach me more on how to prepare such a large volume each time, how to store them so that their quality can be maintained, and if any procedures needed before I can use the stock acrylamide solution (or just take them out and do simple dilution)?
Thx very much.
I used to buy 97% pure acrylamide and purify it.
Wear protective gear. Weigh 800 grams acrylamide and combine with 80 g MB3 or MB6 resin and adjust to 1 liter with Type I water.
Stir 2-3 hours (not overnight) in a 2.8 liter Fernbach flask and large magnetic stir bar. Do not heat. Do not store overnight in presence of resin. Residual catalyst from the resin will polymerize the acrylamide. Filter through a Buchner funnel using Whatman #3 over Whatman #1 into a 4 Liter sidearm flask or use a 0.45 micron disposable filter. Rinse resin. Adjust volume to 1.9 liters. Immediately rinse MB3 resin from Fernbach flask as dry resin will stick very firmly.
Add 40 g bis-acrylamide. Stir till dissolved. (2% bis is a saturated solution.) Adjust volume to 2.0 Liters with Type I water. Aliquot and store at 4°C. Stable > two years.
Clean up the area thoroughly after weighing, dissolving and filtering acrylamide.
(Vournakis, 1981. Gene Amplification and Analysis, Vol. II, p. 280.)
This method results in a > 30-fold savings over buying more highly purified liquid material. Two liters of stock is prepared in order to minimize the number of times the more risky powder must be handled. Can substitute MB1 or MB6 resin. Pharmacia (Isoelectric Focusing, 1982) says MB6 is more efficient.
Thx for your prompt reply.
I just wanna know (1) what is the purpose of further purifying the 97% pure acrylamide using resin, and (2) I have searched online for suppliers for the MB3 or MB6 amberlite resin but i could not find update info on that (probably the name of the amberite resin has changed), is there any existing brands commercially available? and (3) I'd also like to know much deeper on how to rinse the resin, does the purification be performed on ion-exchange column?
THx for ur help, appreciated
1. Some contaminants in low purity acrylamide can cause staining artifacts. Purifying 97% acrylamide was just a way to save money, but it may no longer be available. Use higher purity and simply prepare the 19:1 or 29:1 acrylamide:bis-acrylamide mix and run through a filter.
2. I assume that any Amberlite mixed-bed resin will perform similarly.
3. The resin was rinsed with about 100 mLs of water.
I am new to proteomics and I would like to ask whether in your lab you use (1) powdered form acrylamide or (2) Bio-rad or GE liquid form acrylamide solution to cast gel?
It seems that the powdered form acrylamide as well as bis are far too dangerous to use, although it is much cheaper.
And I'd also like to ask whether you wear a face mask or do in a hood or other important precautions (e.g. when adding TEMED) if you cast a gel.
Thx in advance~
we use powder of 4x crystallized acrylamide; I do not know if such purity is to buy as solution...
We're a quite underfunded institute, but here nobody uses powdered acrylamide. And I wouldn't recommend it to anybody (at least if there are not important reasons for the powder). Luckily nobody uses it very often and no large gels are necessary in our applications. And for this at times uses, acrylamide solutions are the better choice.