Question

Describe the pros and cons of using GFP over chemical fixation and immunofluorescence to study an organelle or protein.

Answer 1

GFP or green fluorescent protein has attained great significance in the field if research, biology, and medicine. The protein is advantageous over chemical fixation as it allows not only the observation but also the movement associated with certain process and other related activities in the cell.

It can be attached to other proteins and can be used in different organisms, the attachment of GFP tag allows the organelle or the protein to be visualized easily. The fusion protein do not interfere in the functioning and location of the protein. But there may be certain disadvantages with smaller proteins as their molecular weight may get affected due to larger molecular weight of GFP hence can interfere sometimes.

These are non toxic and can be used to study the living cell processes, physiology and dynamics. The associated protein's activity is not lost.

Cons: may give background autofluorescence in certain cases. May form insoluble inclusion bodies that may not be fluorescent.

Immunofluorescence uses fluorescent dyes so there is no alteration of molecular weight or formation of inclusion bodies. It provides poor signal than GFP. It is more complex. There may be certain non specific binding of antibody with other antigens.