Dialysis of Colloid Sample
Protocols for today were adopted from Dr. Fox's Notebook.
Description
- Today we dialyzed a 35:1 Au:Protein solution (colloid solution) made earlier and stored in our lab drawer.
- For dialysis, we needed to create 1 L of a 50 mM Glycine buffer, with a pH ~3.5.
- To do this, 4 mL of 1 M HCl and 3.7608 grams of glycine were combined in a 1 L volumetric flask.
- We also needed to prepare 500 mL of a 50 mM NaCl solution.
- To do this, 1.46307 g of NaCl was added to water in a 500 mL volumetric flask.
- Additionally, 25 mL of 1 g/L Lysozyme was prepared.
- To do this, 0.025 g of Lysozyme was combined with water in a 25 mL volumetric flask.
- We then performed dialysis using the protocol described in the "Dialysis Protocol" section below to prepare the dialysis tubes.
- Dialysis was performed by adding 4 mL of the lysozyme solution to:
- A 3500 g/mol MWCO tube in NaCl soak.
- A 3500 g/mol MWCO tube in Glycine Buffer soak.
- A 3500 g/mol MWCO tube in HPLC water soak.
- A 25000 g/mol MWCO tube in Glycine Buffer soak.
- An additional dialysis was performed using 4 mL of the colloid solution in:
- A 3500 g/mol MWCO tube in Glycine Buffer soak.
Dialysis Protocol
- Cut about a 3" length of dialysis tubing
- 25,000 MWCO tubing is stored in an azide solution to prevent mold (use care; NaN3 is toxic)
- keep the 25,000 MWCO tubing wet to prevent pore shrinkage
- wash cut tubing with DI water, both inside and out
- 3,500 MWCO and 15,000 MWCO are dry and need to be wet prior to filling
- 25,000 MWCO needs to be rinsed to remove the sodium azide
- flatten tubing and remove as much residual water as possible with gloves fingers
- clamp one end 1/4 - 1/2" from edge
- open other end and transfer your measured protein solution inside
- carefully flatten open end and clamp, making sure no liquid escapes
- rinse with DI water, particularly the ends, which may have residual protein solution on it
- place into a 150 mL beaker (or 250 mL or 400 mL)
- measure out 100 - 200 mL of your dialyzing solution
- label beaker, cover with parafilm, and leave on shaker on low speed to help prevent gradients during dialysis
|