Day 5 - In Pictures

Today we complete the dialysis! Then move on to freeze-drying!

The Lab: 

 Step 1: DaWei pours out the 5% acetic acid and removed urea 

Step 2: He then replaces it with a fresh solution of 5% acetic acid free of urea! 

Step3: Steps 1-2 are repeated once more at 2-3 hours each time.

Step 4: Placing it in rack and puncturing a hole. This is to ensure that the frozen water and acetic acid can be sublimed when in the freeze dryer, leaving only the protein

It it then placed in a free dryer for 1 h and 30 mins to ensure that everything is frozen.

The -80 Freezer!

Step 5: DaWei quickly places the rack containing the vial in a special glass which can withstand pressures close to vacuum. This has to be done quickly to ensure that the solution does not melt! 

   Done!

This is left overnight to ensure that all water and the 5% acetic is removed, leaving only the protein.

The freeze-dryer machine!

Our attempts successful!:

Day 5 - Completing Dialysis, onto obtaining pure denatured protein

We are halfway through our shadowing programme. Today's procedure is similar to yesterday's.

The dialysis bag containing urea, denatured protein and 5% acetic acid was further processed so that all the urea can removed from the bag. This step was repeated twice, each time roughly 2-3 hours. The result was a bag containing water, 5% acetic acid and the protein, meaning that urea has been successfully removed.

In order to obtain the pure protein powder, the bag was placed in a freezer at -80 degrees Celsius. The bag then turned solid hard. Afterwards, Dawei placed the bag in a freeze dryer. Ice and acetic acid which were initially present soon sublimed. Liquid water and aqueous acetic acid were not  formed as both substances were sublimed. 

The final product contains the protein only. 

Day 4 - Protein Extraction

Pre - Preparation: Protein (Polymer)

DaWei needs to obtain the Unfolded Protein Strand.. First he adds urea to the protein. This urea denaturises the protein into a polymer. After doing so he needs to remove the denatured protein. He then makes adds acetic acid at 5%.

Dialysis:

Step 1: Taking out the right vial containing his prepared solution.

Step 2: The bag to contain the solution is soaked in deionised water!

Step 3: DaWei proceeds to homogenise the 5% acetic acid in deionised (total 3 Litres!) water using a Homogeniser. The white motion blur you see is a magnetic mixer which is spun by a electromagnet below the machine.

Step 4: DaWei Proceeds to carefully add the protein into the bag.

Step 5: Securing the bag with clips!

Step 6: The bag is placed into a container containing roughly 800 mililitres of 5% acetic acid. This first round will be put overnight. Subsequent ones are done for 2-3 hours to remove any excess urea. Stay tuned!

How this last step works: 

The 5% acetic acid is maintained to make sure to make sure the protein stays soluble in the solution. The urea molecules are small enough to pass through the semi-permeable membrane of the bag. This is done repeatedly, removing any remaining urea.

Points to note: Dialysis although share similar steps to osmosis, they are different, to understand do check out a website by clicking here.

Day 4 - More application

Unfortunately my partner has fallen sick, so I will be putting my own thoughts here.

Today's laboratory work is very similar to what I have learnt. They make use of the process of osmosis and in this case, it is termed dialysis. More advanced equipment are used to make the process more effient and make it a much quicker process. Careful measures are also taken to ensure the wanted proteins are not contaminated. Once again, this really lets me appreciate how the practices we do at our level are applied at this higher level. Stay tuned, for further information on the completed work!