Immunoassays probe design

Immunoaffinity chromatography (IAC), 6 400—402 12 137, 145 Immunoanalyzers, automated, 14 150 Immunoassay(s), 14 135-159. See also Immunoassay- DNA probe hybrid assays Immunoassay methods Immuno(bio)sensors antibody-antigen reaction, 14 136-138 basic technology in, 14 138-140 chemiluminescent, 14 150-151 classification of, 14 140-153 design of, 14 139-140 enzyme, 14 143-148 fluorescence, 14 148-150 highly specific, 14 153 historical perspective on, 14 136 microarrays and, 14 156—157 microfluidics in, 26 968—969 monoclonal versus polyclonal antibodies in, 14 152-153... 

Selected entries from Methods in Enzymology [vol, page(s)] Design, 178, 551 immunoassay, 178, 542 production, 178, 531 purification, 178, 543 substrates and enzymatic assay, 178, 544 derivatization with spectroscopic probe, 178, 567 ester cleavage assays, 178, 565 fluorescence quenching binding assay, 178,... 

In general terms, DNA-based assays are easier to develop than immunoassays since the only requirements are the availability of the gene or DNA sequence to be targeted. These sequences are needed to design primers (PCR) and probes (RT-PCR) as well as for determining the G/C ratio to optimize assay conditions. This information can be obtained from databases, and software is available to facilitate the design of primers and probes. These probes have also become less expensive to produce. 

The most significant limitation of immunochemical methods is inadequate sensitivity such that preamplification by PCR is required. Further improvements in solid phases, reporter enzymes, substrates, and hybridization probes will extend the range of analytes amenable to assay without amplification. Solid phases designed specifically for DNA immunoassays are needed. Enzyme engineering for thermostability. 

Amplification with other nanoparticles as ECL labels has also been demonstrated [102-108]. CdSe nanocrystals (NCs) and Qdots have been used for immunoassays. For example, an ECL immunosensor was constructed by immobilizing CdS Qdots and capture antibodies on a poly(diaUyldimethylammonium chloride)-functionalized CNT-modified (PDDA/CNTs) electrode to detect a-fetoprotein [98] A bio-bar-code probe labeled antibody was designed by conjugation of hemin and a single-stranded guanine-rich oligonucleotide to the antibody on gold nanoparticles. The bio-bar-coded probe was captured on the... 

More and better QD probes have been designed and constructed during the past several years, and they are getting closer and closer to clinical applications in the near future. The impact of QDs on immunoassay, diagnostics, and molecular pathology can already be seen. We believe that more robust and reproducible conjugation methods for QD probe production are key for their commercialization and widespread use. Additionally, the discovery of non-tra-ditional QDs will pave the way for the in vivo clinical use of these nanomaterials. 

A large body of very fine work is being performed by flie group of D. Parker at Durham University on luminescent cyclen derivatives with two major aims. One is the design of responsive ratiometric probes for the determination of several analytes, including pH, O2 and various anions within the frame of luminescent time-resolved immunoassays [134]. In this context, a rapid and reliable analytical method enabling the measurement of citrate in seminal fluids has been vahdated citrate is a key metabohte the concentration of which falls by up to a factor of ten in individuals having prostate cancer [135]. 

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