Exceptionally fast uptake and metabolism of cyanidin 3-glucoside by rat kidneys and liver

To asses the hypothesis that anthocyanins are rapidly taken up from the blood into tissues, where they accumulate up to their bioactivity threshold, an intravenous dose of cyanidin 3-glucoside (1) was administered to anaesthetized rats. Cyanidin 3-glucoside (1) and its metabolites were analyzed in the plasma, kidneys, liver, urine, and bile, using last-generation mass spectrometry. Compound 1 was found to rapidly disappear from plasma (t/2=0.36 min). As soon as 15 s after its administration, both 1 and its methylation product, peonidin 3-glucoside (2), were detected in the plasma, kidneys, and liver. At 1 min, both 1 and 2 had almost disappeared from the plasma, but attained their peak concentrations in the kidneys and in the liver. Compound 2 was rapidly excreted both in the bile and in the urine. Three additional methylated metabolites were detected in traces, namely, delphinidin 3-glucoside (3), petunidin 3-glucoside (4), and malvidin 3-glucoside (5). These data contribute to solving the paradox of the high bioactivity of anthocyanins in spite of their apparent low bioavailability.     

Simple and cost-effective isolation of monocytes from buffy coats

We have optimized the procedure of monocyte isolation on a Percoll density gradient. The new procedure consists of three steps: (1) the isolation of MNC on a Ficoll density gradient; (2) the separation of monocytes from lymphocytes on a high-density hyper-osmotic Percoll density gradient; and (3) the separation of monocytes from platelets and dead cells on a low-density iso-osmotic Percoll density gradient. The procedure is simple and cost-effective. Monocyte purity and recovery are both about 75% and platelet contamination is low. The isolated monocytes retain their capacity to differentiate into dendritic cells in vitro.     

Real-time RT-PCR assay for rapid and specific detection of classical swine fever virus: comparison of SYBR Green and TaqMan MGB detection methods using novel MGB probes

Real-time PCR is an accurate, rapid and reliable method that can be used for the detection and also for the quantitation of specific DNA molecules. The basic principle is the recurring measurement of a fluorescent signal, which is proportional to the amount of amplification product. In our trial two detection systems were tested for classical swine fever virus (CSFV) detection and for its discrimination from other pestiviruses; non-specific dsDNA-binding dye SYBR Green and specific fluorogenic TaqMan MGB probes. Real-time RT-PCR assays were evaluated for diagnostic sensitivity and specificity by different pestiviral reference and field strains. With both approaches, SYBR Green and TaqMan probes, respectively, all of the CSFV strains isolated on cell culture were detected and also clearly distinguished from other pestiviruses. However, the established one-step real-time TaqMan RT-PCR assay was shown to be more appropriate for pestivirus quantitation, it reduces the risk of contamination and is less time consuming.