Identifying inhibitors/enhancers of quantitative real-time PCR in food samples using a newly developed synthetic plasmid.

BACKGROUND: Polymerase chain reaction (PCR) has become a common technique offering fast and sensitive analysis of DNA in food/feed samples. However, many substances, either already present in the sample or introduced during sample processing, inhibit PCR and thus underestimate the DNA content. It is therefore necessary to identify PCR inhibition in order to correctly evaluate the sample. RESULTS: We designed and validated a synthetic plasmid DNA that can be used to detect and quantify PCR inhibition. The DNA sequence, appropriate primers and probe, were designed in silico, synthesized and the sequence was inserted into a plasmid vector. The performance of the plasmid was verified via calibration curves and by performing the assay in the presence of various DNAs (crops, fungus, bacterium). The detection of PCR inhibition was assessed using six inhibiting substances with different modes of action, substances used in sample processing (EDTA, ethanol, NaCl, SDS) and food additives (sodium glutamate, tartrazine). The plasmid performance proved to be reproducible and there were no interactions with other DNAs. The plasmid was able to identify the presence of the inhibitors in a wide range of concentrations. CONCLUSION: The presented plasmid DNA is a suitable and inexpensive possibility for evaluating PCR inhibition.

Prevention of contrast agent-induced renal impairment in patients with chronic renal insufficiency and heart disease by high-dose intravenous N-acetylcysteine: a pilot-ministudy.

BACKGROUND: Contrast-induced nephropathy is a relatively common complication occurring after various procedures requiring contrast agent injection, especially in patients with pre-existing renal failure. AIM: This pilot study was designed to assess the effects of a high intravenous dose of N-acetylcysteine (NAC) on plasma creatinine concentration. METHODS: Twenty patients with pre-existing renal insufficiency were given NAC at a dose of 100 mg/kg. No contrast agent was given to 10 patients (Group A), whereas 10 patients received contrast at the time of coronary angiography (Group B). Changes in plasma creatinine were assessed at 3 hours and one day following NAC administration. RESULTS: In Group B, NAC prevented creatinine increase: baseline levels were 210.98+/-77.33 micromol/L, 200.26+/-71.94 micromol/L (NS) after 3 hours, and 203.80+/-83.94 micromol/L 24 hours later (NS). The following was seen in Group A patients: 201.21+/-42.28 micromol/L, 190.31+/-42.74 micromol/L (p<0.01), and 170.08+/-45.53 micromol/L (p<0.01), respectively. CONCLUSION: The results of this study confirm the effectiveness of NAC in prevention of contrast agent-induced renal impairment. In addition, we demonstrated the beneficial effects of NAC on renal function in patients who were not exposed to contrast agent. This pilot study should provide the basis for more comprehensive research and, also, for safe clinical practice.

Successful Treatment of Iron-Overload Cardiomyopathy in Hereditary Hemochromatosis With Deferoxamine and Deferiprone.

There is scarce evidence regarding the use of iron chelators in patients with hereditary hemochromatosis who are intolerant of phlebotomy or erythrocytapheresis. A 52-year-old man with genetically confirmed HFE hemochromatosis presented with liver disease and heart failure with severe left ventricular systolic dysfunction. Because of anemia after initial treatment, we added intravenous deferoxamine followed by oral deferiprone to less frequent erythrocytapheresis, which normalized systolic function within 1 year. Repeated cardiac magnetic resonance imaging revealed improvement of the T2* relaxation time. This report illustrates the beneficial effect of iron chelators in individuals with HFE hemochromatosis and poor tolerance of erythrocytapheresis.