I'm trying to localise a protein called "Protein B."
I've made a vector which encodes protein B that is tagged with an epitope derived from the flu virus. This vector is then transformed into a human cell line, and antibodies to the flu virus will be used to locate the protein.
2 staining procedures are done, one with no alcohol, and one with 2% (v/v) alcohol.
In the staining with no alcohol, you can see uniform staining of the area outside the nucleus.
In the staining with 2% alcohol, you see speckled staining in the area outside the nucleus.
I know that protein B is located in the mitochondria/cytoplasm.
My question is: What does the uniform staining vs speckled staining indicate? Also, why does this type of staining occur?
Uniform staining is for identification of a pattern, and scanning the identity of mitotic or dividing cells. The condensed chromatin of the mitotic cells exhibits solid, uniform fluorescence which is often more pronounced than in the resting cell nuclei.In the uniform pattern, the resting cells exhibit uniform, diffuse fluorescence of the entire nucleus.This characteristic pattern is often the result of anti- B proteins.
Speckled staining is for the purpose of a speckled pattern. In speckled pattern, the mitotic cells show no staining of the condensed chromosomal regions. The resting cells exhibit grandular fluorescence through out the entire nucleus. The nuclear speckling can be defined as coarse or fine. The Coarse speckling pattern is often the result of anti- B protein.