Dot blot

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Schematic of the use of two ASO probes on duplicate dot blot filters.

A dot blot (or slot blot) is a technique in molecular biology used to detect biomolecules, and for detecting, analyzing, and identifying proteins. It represents a simplification of the northern blot, Southern blot, or western blot methods. In a dot blot the biomolecules to be detected are not first separated by electrophoresis. Instead, a mixture containing the molecule to be detected is applied directly on a membrane as a dot, and then is spotted through circular templates directly onto the membrane or paper substrate. This differs from the western blot because protein samples are not separated electrophoretically. This is then followed by detection by either nucleotide probes (for a northern blot and southern blot) or antibodies (for a western blot).

The technique offers significant savings in time, as chromatography or gel electrophoresis, and the complex blotting procedures for the gel are not required. However, it offers no information on the size of the target biomolecule. Furthermore, if two molecules of different sizes are detected, they will still appear as a single dot. Dot blots therefore can only confirm the presence or absence of a biomolecule or biomolecules which can be detected by the DNA probes or the antibody.

A radioactive sample can be hybridized to allow for the detection of variation between samples. The DNA is quantified and equal amounts are aliquoted into tubes in excess of the number of its targets in the samples, such as 10 µg for a plasmid and 1 µg for a PCR amplicon. These are denatured (NaOH and 95°C) and added to the wells where a vacuum sucks the water (with NaOH and NH4OAc) from underneath the membrane (nylon or nitrocellulose).

What can this test detect?[edit]

The sensitive dot blot test can be used to detect the Chlamydia trachomatis infection and other sexually transmitted diseases. Dot blot is used to detect Antidiacyltrehalose Antibodies in Tuberculous patients and Typhoid Fever. This test could increase the number of lives saved that are affected with these diseases. Using the dot blot test can be useful for under-developed countries especially. The dot blot is a good positive predictor of these diseases for countries and regions lacking in medical facilities and laboratories. By using this test many lives could be saved and a cure could be found for the types of diseases this test can detect[citation needed].

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