Electrophoresis Articles

Autoradiography is the use of X-ray (or occasionally photographic) film to detect radioactive materials. It produces a permanent record of the positions and relative intensities of radiolabeled bands in a gel or blot. Typically, biomolecules are labeled with 32P or 35S, and detected by overnight film exposure. The table below gives the amounts of commonly…

One example of one might attempt alkaline blotting is listed here. Alkaline Blotting: The Procedure Positively charged nylon membranes allow the use of alkaline transfer buffers, which link the nucleic acids to the membrane without UV crosslinking. In some cases, alkaline blotting gives a higher background. Increasing the concentration of blocking reagent will often eliminate…

One possible procedure for Southern blotting can be found here. The Procedure The protocols given are for Genomic Southern Blotting, one of the most sensitive Southern techniques. This set of protocols will give superior results in any standard Southern application. SAMPLE PREPARATION Genomic DNA samples must be extremely pure,with the A260/A280 ratio being equal to…

The Procedure ELECTROPHORETIC SEPARATION RNA samples should be free of contaminating particles and DNA, with an A260/A280 ratio of 1.9-2.0. For highly expressed genes, load 10µg total of RNA. For trace expression detection, load 10µg of poly A+ RNA. Ethidium may be added to the gel (4 µl of 10 mg/ml per 100ml gel =…

A DNA or RNA probe will selectively hybridize with nucleic acid molecules of complementary sequence in a sample. A labeled nucleic acid molecule of known sequence can facilitate detection of any complementary molecules in an unknown sample. This is the basis of the RNase protection assay, and PCR amplification, among other techniques. In theory, labeled…

Silver staining is more sensitive than ethidium bromide for double stranded DNA, and detects single stranded DNA or RNA with no loss in sensitivity. Silver staining relies on the reduction of silver cations to insoluble silver metal by nucleic acids. This chemical reaction is insensitive to the macrostructure of the DNA molecule. Reduced silver grains…

National Diagnostics’ Nuclistain offers a significant increase in sensitivity over UV shadowing, along with the convenience of visual staining. With Nuclistain there is no need for UV light. Nuclistain is a blue dye that binds to DNA, revealing blue bands after destaining with water. Nuclistain detects DNA down to levels of 50ng. Note: Nuclistain does not…

When ultraviolet light strikes a DNA molecule, it may be absorbed, transmitted, or, if a fluorescent dye is present, re-emitted as visible light. Detection of DNA/RNA in solution is generally done by measuring its UV absorbance at 260 nm. This absorbance is due to the ring systems in the nitrogen bases, and can also be…