Gels and membranes

Proteins and nucleic acids can be separated within gels

Experimental biology depends on two principles of molecular recognition that underlie the specificity of assay detection:

1. 1. Linear nucleic acids are identified by sequence-specific hybridization with synthetic complementary DNA sequences (oligonucleotide probes).

2. 2. Three-dimensional or denatured protein structures are identified by epitope-specific antibody binding.

Most assays detecting these phenomena involve the electrophoretic separation of molecules using porous jelly-like slabs termed gels. Indeed, for the last two decades, many scientific manuscripts have contained few data other than those involving visualization of molecules within gels. There are many different gel compositions, with the most common being agarose gels (used for separating nucleic acids) and polyacrylamide gels (used for separating proteins).

Gels are modified for specific purposes; for example, formaldehyde is often added to urea-containing agarose gels when RNA is being electrophoresed. Nucleic acids can be visualized on agarose gels by adding the intercalating agent ethidium bromide, which fluoresces on absorbing ultraviolet light. Similarly, the detergent sodium dodecyl sulfate (SDS) is added to protein gels when polypeptide denaturation is desired – denaturation destroys the noncovalent secondary structure of proteins, permitting linearization and hence more accurate electrophoretic measurement of molecular weight.