OBJECTIVEThis study compared two newborn Cytomegalovirus (CMV) IgG antibody screening ELISA kits and evaluated the detection effectiveness of Abnova kit.

METHODSCMV IgG antibodies were detected by both SeraQuest and Abnova kits from dried blood spot (DBS) samples of 488 newborn heel sticks. The detection abilities of these two kits were compared in different sample dilution concentrations. Relative detection effectiveness of the Abnova kit was defined by statistical method using the SeraQuest kit as a point of comparison.

RESULTCompared to the SeraQuest screening test kit, the Abnova kit revealed a sensitivity of 98.9%, specificity of 78.6%, positive predictive value of 99.3%, negative predictive value of 68.8%, and the coincidence rate for these two screening test kits at 98.3%. The consistency check of both kits based on interpretation of the kappa statistic was relatively good. For the Abnova kit, the "area under the ROC curve" was 0.887, which indicates moderate accuracy.

CONCLUSIONAbnova kit can be applied to newborn screening for congenital CMV infections. However, repeating the test for ambiguous results is suggested to increase the specificity and negative predictive value.

Antibodies, Viral ; blood ; Cytomegalovirus ; immunology ; isolation & purification ; Cytomegalovirus Infections ; blood ; diagnosis ; virology ; Enzyme-Linked Immunosorbent Assay ; instrumentation ; methods ; Female ; Humans ; Immunoglobulin G ; blood ; Infant ; Infant, Newborn ; Male ; Neonatal Screening ; methods ; Reagent Kits, Diagnostic

OBJECTIVETraditional detection approaches for non-O157 STEC are both time and labour consuming in diseases surveillance. Virulence genes detection based on multiplex PCR could not only improve the detection efficiency but also increase the accuracy.

METHODSSix virulence genes of non-O157:H7 (stx1, stx2, eae, hly, etpD, katP6) were detected by two groups of trebling PCRs. The multiplex PCRs were optimized by melting curve analysis in SYBR Green I real-time PCR. Testing result of multiplex PCR was consistent with serological testing.

RESULTSThe sensitivity limits of the multiplex PCR for stx1, stx2, eaeP, etpD, katP, and hly were 10 ng/ml, 120 ng/ml, 110 ng/ml,165 ng/ml, 85 ng/ml, and 15 ng/ml, respectively, which is similar with that of single PCR. When the multiplex PCR was applied in 120 adults and 90 children diarrhea samples detection, 13 cases were detected for non-O157 positive.

CONCLUSIONThe method we established can be used for non-O157 STEC virulence genes detection and screening with high efficiency and accuracy.

Escherichia coli Infections ; diagnosis ; microbiology ; Escherichia coli Proteins ; genetics ; Humans ; Multiplex Polymerase Chain Reaction ; methods ; Shiga-Toxigenic Escherichia coli ; genetics ; isolation & purification ; Virulence Factors ; genetics

OBJECTIVETo establish chemiluminescence enzyme immunoassay (CLEIA) for quantitative detection of procollagen III N-terminal peptide (P III NP) in serum.

METHODSA sandwich reaction was preformed with horseradish peroxidase labeled monoclonal antibody of P III NP as the catalytic enzyme and the luminol as the luminescence reagent. Several reactions liquid's concentration and reaction conditions were optimized. The method was evaluated in all aspects such as linear range, sensitivity, specificity, stability and so on. The CLEIA was compared with imported ELISA kits, by detecting clinical serum.

RESULTSThe linear range was 0.8-85 ng/ml. The detection limit was 0.5 ng/ml. Inter-assay and intra-assay RSD were both less than 10%. The recoveries of three different spiked concentration samples were 96.2%, 91.2% and 101.1%. After stored at 4 degrees C and 37 degrees C for 3, 5, 7 days, the analysis showed correlation coefficient higher than 0.99 and RSD lower than 6%. The detected results of clinical sera with CLEIA closely corresponded to those with imported ELISA.

CONCLUSIONEstablished CLEIA for quantity determination of serum P III NP has high accuracy, sensitivity and repeatability.

Adult ; Aged ; Antibodies, Monoclonal ; analysis ; Enzyme-Linked Immunosorbent Assay ; instrumentation ; methods ; Female ; Humans ; Liver Cirrhosis ; blood ; diagnosis ; Luminescence ; Luminescent Measurements ; methods ; Male ; Middle Aged ; Peptide Fragments ; blood ; chemistry ; Procollagen ; blood ; chemistry ; Sensitivity and Specificity

OBJECTIVETo establish enzyme-linked immunosorbent assay (ELISA) for quantitative detection of Golgi protein73 (GP73) in serum.

METHODSA sandwich reaction was preformed with horseradish peroxidase labeled monoclonal antibody of GP73 as the catalytic enzyme. Several reactions liquid's concentration and reaction conditions were optimized. The method was evaluated in all aspects such as linear range, sensitivity, specificity, stability and so on.

RESULTSThe linear range was 25-500 ng/ml. The detection limit was 18.5 ng/ml. Inter-assay and intra-assay RSD were both less than 10%. The recoveries of three different spiked concentration samples were 95.3%, 92.6% and 103.7%. After stored at 4 degrees C and 37 degrees C for 3, 5, 7 days, the analysis showed correlation coefficient higher than 0.98 and RSD lower than 10%.

CONCLUSIONEstablished ELISA for quantity determination of serum GP73 has high accuracy, sensitivity and repeatability.

Antibodies, Monoclonal ; analysis ; Enzyme-Linked Immunosorbent Assay ; methods ; Humans ; Membrane Proteins ; blood ; Sensitivity and Specificity

OBJECTIVEThe purpose of this study was to develop RT- PCR assay for Rapidly detect and distinguish between Norovirus genogroup I and genogroup II with a pair of primers.

METHODSA pairs of primers specific to capsid prote in ORF2 gene of G I and G II Norovirus were dsigned according to the published complete genome sequence, with which the RNA of Norovirus was extracted and RT-PCR amplification. The sensitivity, specificity of the RT- PCR assay was estimated and apply it to the detection of Norovirus in clinical specimens.

RESULTSThe results showed that the assay possessed high specificity for Norovirus detection and without any evident cross-reaction with other viruses, including rotavirus, enteric adenovirus and hepatitis A virus. The detection limit of RT-PCR assay for Norovirus G I and G II were up to 100 pg/ml and 10 pg/ml respectively.

CONCLUSIONThe RT- PCR assay provide rapid and sensitive detection of Norovirus G I and G II and should prove to be useful for Norovirus diagnosis in the outbreaks of acute gastroenteritis.

Caliciviridae Infections ; diagnosis ; virology ; DNA Primers ; genetics ; Gastroenteritis ; diagnosis ; virology ; Humans ; Norovirus ; classification ; genetics ; isolation & purification ; Reverse Transcriptase Polymerase Chain Reaction ; instrumentation ; methods

OBJECTIVETo establish a localization ultrathin section method through which target cytopathic cells could be sectioned in situ.

METHODSLab-Tek Chamber slide system (177402) was selected as resin embedding mould. Cells infected with Human adenovirus type 5 (Ad5) or A/HN/SWL3/ 2009 (H1N1) influenza virus were embedded in situ as models. Target cytopathic cells were exposed by trimming, sectioned and observed under transmission electron microscope (TEM).

RESULTSTarget cells could be sectioned in situ and virus particles could be found easily on sections.

CONCLUSIONA localization ultrathin sectioning method was established and this technique could be applied in virus detection in cytopathic cells to improve TEM detection efficiency.

Adenovirus Infections, Human ; pathology ; virology ; Adenoviruses, Human ; physiology ; ultrastructure ; Cell Line ; Humans ; Influenza A Virus, H1N1 Subtype ; physiology ; ultrastructure ; Influenza, Human ; pathology ; virology ; Microscopy, Electron, Transmission ; Microtomy ; methods

OBJECTIVETo establish a method to produce virus-like particles (VLP) of poliovirus type I in Saccharomy cescerevisiae to develop potential novel recombinant vaccine against poliovirus type 1.

METHODSThe genes of P1 and 3CD of poliovirus type I were optimized, synthesized and inserted into expression vector, which was further transfected into Saccharomy cescerevisiae. The extracts of yeast cells were purified by CsCl density gradient centrifugation after induction and cell lysis.

RESULTSElectrophoresis and sequencing analyses showed that the genes P1 and 3CD of poliovirus type I were successfully inserted into expression vector and encode a protein whose amino acid sequences were identical with wide-type genes of poliovirus type I. Electronic microscopy analysis showed that the VLPs of poliovirus type I could be efficiently formed in Saccharomy cescerevisiae.

CONCLUSIONThe VLPs of poliovirus type I could be efficiently produced by co-expression of P1 and 3CD genes in Saccharomy cescerevisiae.

Female ; Gene Expression ; Humans ; Male ; Poliomyelitis ; prevention & control ; virology ; Poliovirus ; genetics ; metabolism ; Poliovirus Vaccines ; genetics ; metabolism ; Saccharomyces cerevisiae ; genetics ; metabolism ; Viral Proteins ; genetics ; metabolism ; Virion ; genetics ; metabolism

OBJECTIVETo study cellular and humoral immune status on prophase of severe hepatitis B (PSHB).

METHODS56 cases of PSHB patients, 40 cases of chronic hepatitis B (CHB) patients and 20 cases of healthy volunteers were enrolled for detection of CD3+, CD4+, CD8+ and CD3-/CD19+ (B cells) lymphocyte subsets in peripheral blood by flow cytometry. Serum IgG and complement C3 was detected by immunoturbidimetry and analyzed statistically.

RESULTSCompared with CHB group and healthy control group, percentage of lymphocyte subsets CD8+ were significantly lower in PSHB group (P < 0.01 or P < 0.05). While the percentage of lymphocyte subsets CD4+ and ratio of CD4+/CD8+ in PSHB group was obviously higher than those in CHB group (P < 0.+01 or P < 0.05). In addition, There was no significant difference on the percentage of B cell and level of serum IgG between PSHB group and CHB group (P > 0.05, while the level of serum complement C3 in PSHB group were significantly lower than those in CHB group and healthy control group (P < 0.01, P < 0.05).

CONCLUSIONPSHB has a certain degree of cellular immune dysfunction, which characterized by cellular immune function hyperfunction and humoral immune suppression.

Adult ; Antibodies, Viral ; immunology ; Complement C3 ; immunology ; Female ; Flow Cytometry ; Hepatitis B, Chronic ; immunology ; Humans ; Lymphocyte Count ; Male ; Middle Aged ; T-Lymphocyte Subsets ; cytology ; immunology ; Young Adult

OBJECTIVEA retrospective study was conducted to investigate the clinical features and prognostic factors of 73 cases of severe hepatitis.

METHODSTo summarize clinical features of 73 cases of severe hepatitis, grouping by etiology and pathogenesis. A retrospective analysis was performed to evaluate the relationship between biochemical characteristics (liver function, renal function, electrolytes, PTA, etc) and complications (hepatic encephalopathy, upper gastrointestinal bleeding, hepatorenal syndrome, ascites, abdominal infections, etc) and prognosis.

RESULTS(1) HBV infection alone accounted for 65.75%. Alcoholic liver disease, drug-induced liver injury, hepatitis E, autoimmune hepatitis, overlapping causes and other factors were five cases (6.85%), six cases (8.22%), two cases (2.74%), two cases (2.74%), seven cases (9.59%) and three cases (4.11%) respectively. According to the incidence rate, severity and underlying liver condition, subacute hepatitis, cases based on chronic hepatitis and on cirrhosis were 12 cases (16.43%), 11 cases (15.07%), 50 cases (68.49%) respectively. Clinical manifestations with or without hepatic encephalopathy accounted for 58.90% or 41.10%. (2) The highest mortality of severe hepatitis was alcoholic liver disease and patients on the basis of overlapping factors (66.67%), followed by autoimmune liver disease (50%). The mortality of HBV-related hepatitis was 18.75%. Overall mortality of 73 cases of severe hepatitis was 28.77%, of which cirrhosis group was higher than non-cirrhotic group (40% vs 4.3%, P = 0.002). The difference was statistically significant. Patients without hepatic encephalopathy had lower mortality than with hepatic encephalopathy (3.33% vs 46.51%). The mortality of patients with hepatic encephalopathy Stage III and IV was 72.73%. (3) Independent samples t test filtered nine factors associated with death, namely cirrhosis, upper gastrointestinal bleeding, hepatic encephalopathy, hepatorenal syndrome, serum creatinine, total bilirubin (TBIL), direct bilirubin (DBIL), albumin (ALB) and serum sodium. The results of multivariate conditional logistic regression analysis indicated that hepatic encephalopathy, serum creatinine levels were risk factors for death, whereas ALB as a protective factor.

CONCLUSIONHepatic encephalopathy, serum creatinine levels were risk factors for severe hepatitis death, But ALB was protective factor. Nucleotide analogs using was the main reason why the mortality of hepatitis B was as low as 18.75%.

Adult ; Aged ; Female ; Hepatitis ; complications ; mortality ; pathology ; virology ; Hepatitis B virus ; genetics ; isolation & purification ; Humans ; Male ; Middle Aged ; Prognosis ; Retrospective Studies ; Risk Factors

OBJECTIVETo study the efficacy of early, enough methylprednisone combined with immunoglobulin in treatments of severe hand-foot-mouth disease.

METHODS568 patients of severe hand-foot-mouth disease involved were randomized into group A and group B. Patients of both groups all accepted symptomatic treatment, supportive therapy and the treatment of control intracranial pressure. And patients in group A received the treatment of early, enough methylprednisone combined with immunoglobulin, whereas patients in group B received conventional therapy. RESULTS; Curative rate of group A was better than that of group B, otherwise incidence rate of critical illness was less than that of group B. The control time of fever, erythra, neurological symptoms, features of pneumonia and increased leukocyte of group A was shorter than that of group B, and no more recent or Long-term treatment-related adverse reaction in group A.

CONCLUSIONEarly, enough methylprednisone combined with immunoglobulin is effective, safe in treating severe hand-foot-mouth disease, and worthy to recommand its clinical use.

Adult ; Aged ; Biomedical Research ; Drug Therapy, Combination ; Female ; Hand, Foot and Mouth Disease ; drug therapy ; Humans ; Immunoglobulins ; therapeutic use ; Male ; Methylprednisolone Hemisuccinate ; therapeutic use ; Middle Aged