Question

You raise a monoclonal antibody to a protein that is highly expressed in germinating pollen grains of Brassica napus. Outline the procedure that you would use to identify the location of this protein within cells.

Answer 1

Immunofluorescence and immunohistochemistry

These are the methods that are used for protein detection.in this method antibodies are used for detection and localization of proteins and other antigens within biological samples. Tissues and other samples are prepared by a process call “fixation” which is usually done directly on microscopy slides. As a result, cells or tissues retain cellular antigen distribution and preserve its cellular morphology. Before the detection, samples are subjected to a permeabilizing process exposing the antigens which are usually not accessible, thus allowing the antigen detection within the structures.

These methods are widely used in autoimmune disease research and diagnostics. They are especially useful to identify structures to which antibodies are bound within the fixed cells. In addition, these methods allow identification of regions within cells that are important for binding of proteins or specific antigens.

Other methods such as western blotting, Elisa, mass spectrometry can also be used.