Competitive binding methods, alternative

Alternatively, MIPs have also been used in biological receptors for competitive binding assays. The assay principle is similar to that in other known biological assays such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA) except that instead of antibodies, MIPs are utilized. This method is often called molecularly imprinted assay (MIA). Typically, in MIA methods, a marker molecule (a labeled analyte analogue) is incubated together with the sample and the MIPs. Analyte and marker molecules compete for the binding... 

Once the RNA is prepared, it is converted to cDNA and labeled either with radioactive precursors ([33P]-dCTP or [32P]-dCTP) or with non-radioactive fluorescent labels such as Cy3-dNTP and Cy5-dNTP. Radioactive labels are commonly used for membrane approaches while fluorescent probes are the method of choice for glass slides. When using fluorescent probes, cDNA prepared from controls can be labeled with one dye while cDNA from treated samples can be labeled with the other. These are then mixed and used in competitive binding to probes on the chip. Alternatively, one can compare all control and experimental samples to the same reference RNA sample, labeled with one of the dyes. Reference RNA samples are useful for multiple sample analysis and to account for chip-to-chip variations. In the case of the Affymetrix technique, cDNAs are converted back into fluorescently labeled RNA targets, which bind more tightly than cDNA targets to the short oligonucleotides present on those chips, and the control and experimental samples are applied individually to separate chips. 

The number of disadvantages outweigh the advantages of radioimmunoassays and this leads to development of non-radioisotopic immunoassay methods. Many substances can be firmly attached to the antibody without changes in binding properties towards the antigen. This principle has been used in some methods to be competitive with radioisotopic methods. In order for alternate methods to function as alternatives to radioisotopic... 

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