Enzyme Linked Oligosorbent Assay ELOSA

ELOSA, which is a typical technique equivalent to enzyme-linked immunosorbent assay (ELISA), permits specific capture and detection of a target DNA or RNA sequence. The principle of this technique is shown in Fig. 25a for flat solid support and Fig. 25b for colloidal particles. A capture ODN (ODN-1) with an appropriate sequence is immobilized on a solid support. This ODN specifically captures the complementary single-stranded nucleic acid (DNA or RNA) via hybridization process. To point out the specific capture of the target, a second ODN-bearing HRP enzyme (ODN-2) is then hybridized on a given part of the target. The quantification is then performed via enzyme oxidation leading to a colored or fluorescent product. The fluorescence intensity is proportional to the captured target concentration (Fig. 25b).

Major research in biomedical diagnostic has consisted of increasing the capture and the detection of nucleic acid targets without any polymerase chain

FIG. 25 Enzyme-linked oligosorbent assay (ELOSA). (a) Basic system (ODN probe 1 adsorbed on the plate). (b) Modified ELOSA procedure (latex/ODN probe 1 conjugate adsorbed to the plate).

reaction (PCR) or reverse transcriptase polymerase chain reaction (RT-PCR) amplification step. The efficiency of latex-ODN conjugate in ELOSA has been recently validated using an automated diagnostic apparatus (Vidas from bioMer-ieux S.A.) [23]. Such methodology has been derived from the interesting results obtained using linear polymers on which single-stranded DNA fragments were chemically grafted to amplify the sensitivity of ELOSA [24].

Figure 26 shows some results obtained using latex-ODN conjugate in ELOSA test in which the amplified signal from enzymatic reaction of the ODN/ latex particle ratio. The latex-ODN conjugates are adsorbed onto the solid support of the automated system; then the capture and detection of a model nucleic acid target are investigated as illustrated in Fig. 25b. The sensitivity result observed using latex-ODN conjugate was appreciable and was enhanced by combining reactive polymers/latex particles/ODN in both capture and detection steps [23,25].

FIG. 26 Evaluation of two-dimensional latex assemblies obtained with covalent conjugates (♦) and conjugates obtained by ODN adsorption (•). (From Ref. 23.)

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