RESUMO
Newly developed light-up probes offer an attractive tool for PCR product detection. The light-up probe, which consists of a thiazole orange derivative linked to a peptide nucleic acid oligomer, hybridizes specifically to complementary nucleic acids. Upon hybridization the thiazole orange moiety interacts with the nucleic acid bases and the probe becomes brightly fluorescent. This eliminates the need to separate bound from unbound probes and reduces the risk of cross contamination during sample handling. We demonstrate here the applicability of light-up probes in two different PCR assays, one directed towards the human beta-actin gene and the other towards the invA gene of Salmonella. The probes do not interfere with the PCR reaction and can either be included in the sample mixture or added after completed amplification. The specificity of the probe is found to be excellent: a single-base mismatch in the target sequence is sufficient to prevent probe binding as indicated by the lack of fluorescence increase. Furthermore, a clear correlation is found between the intensity of gel bands and the measured probe fluorescence in solution, which suggests that the amount of PCR products can be quantified using light-up probes.
Assuntos
Sondas de DNA/química , Reação em Cadeia da Polimerase/métodos , Actinas/genética , Proteínas de Bactérias/genética , Pareamento de Bases , Benzotiazóis , Corantes Fluorescentes/química , Humanos , Ácidos Nucleicos Peptídicos/química , Quinolinas , Salmonella/genética , Sensibilidade e Especificidade , Tiazóis/químicaRESUMO
Fifty patients, with single-level disc herniations verified by computed tomography, took part in a study where they were randomized to either microsurgery or standard surgery. The aim of the study was to describe the changes on a series of radiographic examinations after disc resection, and to relate these changes to clinical symptoms. The study was prospective and the patients were examined with contrast-enhanced computed tomography during the first postoperative week, after 1-2 months, and after 1 year. At 1 year after surgery 16 patients showed posterior disc protrusion, 47 showed scar tissue, and 13 showed nerve root displacement. Microsurgically operated patients did not show less scar tissue. None of the postoperative radiographic changes had any definite correlation to remaining back pain or leg pain.
