Question

In a brief paragraph, explain how soluble proteins were obtained from fish muscle samples. Your explanation should include:

1. the purpose of mincing (cutting) and vortexing the tissue [2 pts,]
2. the purpose of the homogenization buffer [2 pts,] and
3. the purpose of the centrifugation step [2 pts.]
You must clearly describe what role each of these played in extracting and solubilizing protein from the samples to receive full credit. Revisit the "Micropipetting and Preparing Solutions" exercise to remind yourself of what the ingredients in the homogenization buffer were.

Answer 1

Step-by-step explanation:

This question is a practical question, however the processes involved in obtaining proteins after "sacrifice" is similar.

After removing the desired muscle sample from the fish during sacrifice, the muscle sample can be stored temporarily in a phosphate buffer saline which can also help remove blood and some other contaminants around the muscle sample. The sample can then be removed and weighed with the desired mass cut into small pieces in order to achieve a smaller surface area which assists for easy homogenization and buffer interaction. The homogenization buffer (which can be a phosphate buffer) is added to this small pieces and then "vortexed" so that the buffer can interact properly with the pieces. The purpose of the homogenization buffer is to prevent osmotic damage in the muscle cells while maintaining a constant pH (depending on the protein to be obtained). After homogenization with an homogenizer, the muscle cell membrane is ruptured and the soluble proteins dissolves into homogenization buffer (which must also be soluble to the desired protein) with a mass of suspended muscle tissues also in the buffer. This mixture is then centrifuged so as to obtain the dissolved/soluble protein (in the homogenization buffer) and separate it from the suspended masses in the mixture.