Approaches to Detecting Proteins by Fluorescence Microscopy

Approach #1:

Immunofluorescence using fluorescently-labeled antibody that binds to protein of interest

secondary antibody can be used to enhance primary antibody

Approach #1: steps

1. Fix the sample

2. Permeabilize the sample

3. Add antibodies

4. Detect using fluorescence microscopy

Approach #2

Immunofluorescence using

fluorescently-labeled antibody binds to a protein tag that’s attached to protein of interest

Generating a Fusion Protein

1. DNA encoding protein of interest

2. DNA encoding protein tag covalently attached to the DNA encoding protein of interest

3. transcription

4. translation

Approach #3:

Direct fluorescence of a fluorescent protein tag attached to the protein of interest

Summary

#1 Immunofluorescence using an antibody that binds to the protein of interest

#2 Immunofluorescence using an antibody that binds to a protein tag attached to the protein of interest

#3 Direct fluorescence of a fluorescent protein attached to the protein of interest