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ObjectiveTo assess the clinical application value of saliva samples from patients with herpes zosterzoster.MethodsSaliva and serum samples of 55 patients with herpes zoster were collected. Patients were split into mild and moderate group and serious group based on the clinical score table. Paid a follow-up visit for the occurrence of PHN three months after the lesion healed. Real-time fluorescence quantitative PCR was used to detect varicella-zoster virus load in saliva and serum, and then observed and verified by transmission electron microscope.ResultsThe positive rates of VZV DNA in saliva and blood samples were 87% and 85% respectively. Transmission electron microscopy showed virus pellets in saliva of patients. The VZV DNA load in saliva in serious group (5.79±2.94) copies/mLwas overtopped than that in mild to moderate group (3.06±2.59) copies/mL (t=3.58, P=0.00). Compared the viral loads in saliva of PHN group(5.82±3.12) copies/mL and non-PHN group (3.84±2.96) copies/mL, t=1.54, P=0.13.ConclusionThe positive rate of VZV DNA in saliva of herpes zoster patients is high, the load is related to clinical symptoms. So saliva can be used for the diagnosis of HZ and assessing the patients′ condition.
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<p><b>OBJECTIVE</b>To prepare oligo microarrays for hepatitis virus detection and genotyping.</p><p><b>METHODS</b>By analyzing the DNA or cDNA of HBV, HDV and 4 different genotypes of HCV with the BLAST program, a group of specific sequences for the candidate probes was specified. Array Designer 3.0 software was applied to analyze the candidates to select probes with high specificity, identical length and similar melting temperature (Tm). Altogether 16, 8 and 68 oligonucleotide probes were designed for diagnosis of HBV, HDV, and genotyping HCV. Following the synthesizing and purification, oligo probes were deposited on oligonucleotide chips as microarrays for hepatitis virus detection and genotyping. The samples were labeled by RD-PCR method. Hybridization results were analyzed to cross out those probes with low specificity and sensitivity, and those with signal to noise ratios (SNR) less than 4.0.</p><p><b>RESULTS</b>Two types of gene chips were successfully developed: microarrays for HBV and HDV simultaneous detection and for HCV genotyping.</p><p><b>CONCLUSION</b>Using oligo probes to construct gene chips for clinical diagnosis of hepatitis virus is a simple and effective method. It may be widely used in detecting hepatitis viruses and their genotyping in clinical settings.</p>
Subject(s)
Base Sequence , DNA Fingerprinting , DNA, Viral , Genetics , Genotype , Hepatitis B virus , Classification , Genetics , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis , Oligonucleotide Probes , Sensitivity and SpecificityABSTRACT
To construct and express Hsp70-HSV2gD fusion protein. Genes of Hsp70 and HSV-2gD were subcloned into vectors pGEX-4T-1 respectively. After confirmed by DNA sequence analysis, the recombinant plasmids pGEX-4T-HSP-gD was transformed into E. coli DH5alpha and induced to express with IPTG. The expressed protein was characterized by SDS-PAGE and Western blot after purified. BALB/c mice were immunized with fusion proteins respectively via intra-m uscular injection. The proliferation of spleen lymphocytes, the level of y-IFN in culture and anti-HSV-2gD IgG antibody in serum was detected was detected. The expressed protein was analyzed by SDS-PAGE after induced with IPTG, which showed a new band with an apparent molecular mass corresponding to the predicted size (118 kD). Western Blotting analysis demonstrates that the purified Hsp70-HSV2gD fusion protein had specific binding activity. The stimulation indexes of spleen lymphocytes, the level of gamma-IFN in culture and anti-HSV-2gD IgG antibody in serum of GST-Hsp70-gD group was obviously higher than that of other groups (P < 0.05 respectively). The successful expression of the Hsp70-HSV2gD fusion protein, which can induce immune responses, laid a solid foundation for its further research.