Methodological aspects of Universal immuno-PCR on standard tubes

ABUD, J.E.; RODRIGUEZ, H.A.; OGGERO, M.; SANTAMARÍA, C.G.

ANALYTICAL BIOCHEMISTRY

ACADEMIC PRESS INC ELSEVIER SCIENCE

Año: 2019 vol. 570 p. 56 - 61

0003-2697

One of the most used formats in inmuno-polymerase chain reaction (IPCR) is known as ?Universal? IPCR (signalgenerating complexes is based on conjugates of biotinylated DNA, biotinylated IgG and avidin). In the present study, we evaluated the utility of using mono- and bi-biotinylated DNA probes, pre-self-assembled DNA-neutravidin complex, blocking step and glutaraldehyde pretreatment of standard PCR tubes to improve the analytical performance of the hTSH-IPCR assay. The use of pre-self-assembled mono-biotinylated DNA-neutravidin complex enhances both the sensitivity and the reproducibility of the hTSH-IPCR assay, even without blocking step: hTSH-IPCR assay showed an improved limit of detection (LOD: 0.01 μIU/ml), calibration sensitivity (SEN: 2.4) and analytic sensitivity (γ: 9 μIU/ml−1) in comparison with both a self-made ELISA and a commercial one.