Purification, characterization, and identification of proteins
or nucleic acids often require analysis of the target in blotting
applications. Proteins or nucleic acids are usually separated by
size using gel electrophoresis and transferred to a membrane
with a blotting technique.
A specific protein can be identified from a mixture using
different detection methods. Primary antibodies can be used
to recognize unique epitopes or genetically engineered fusion
protein tags. Lectins with specificities to certain carbohydrate
residues can be used to probe for a particular glycoprotein.
These reagents can then be detected and visualized with an
enzymatic or fluorescent detection method.
Nucleic acid blotting applications also require high specificity
which is derived from the uniqueness of the target sequence
and the fidelity of the complementary probe. The target
sequence is detected by a complementary probe labeled with
tags such as biotin, digoxigenin (DIG), DNP, or fluorescein.
The label is then detected with an antibody to the tag and
subsequent detection reagents.
Detection sensitivity for both protein and nucleic acid blotting
applications depends on the abundance of the target and
the quality of the reagents available for detecting the primary
antibody or hybridized probe.