An open-reading frame designated 1R1, on DNA component 1 of bean golden mosaic geminivirus (BGMV), has been identified as the coat protein gene. A DraI restriction fragment of BGMV DNA1 that includes ...1R1 was inserted into the SmaI pKK223-3 expression vector. The 32 kD protein expressed in Escherichia coli cells reacted with antibodies to the BGMV capsid polypeptide and behaved identically to purified capsid protein in western blots.
As the cDNAs encoding A
1aB
1b and A
2B
1a subunit precursors of the glycinin A
2 subfamily contain a unique
NcoI site sequence, (A)CCATGG, occurring at their translation initiation sites, plasmids ...were constructed to direct the synthesis of those precursor proteins by inserting
NcoI/
PstI fragments derived from those cDNA clones into the
Nco/
PstI-pKK233-2 expression vector in
Escherichia coli MV 1190, respectively. The resultant plasmids directed the expression of 57-kDa protein components that have molecular masses in agreement with those of the in vitro translation products directed by glycinin A
2 subfamily mRNAs, by the addition of isopropyl β-D-thiogalactoside. These proteins, which comprised as much as 1% of the total bacterial protein, are immunoprecipitable with rabbit antibodies specific for glycinin subunits. This procedure makes glycinin subunits available as a model for studying structure-function relationships in seed proteins using site-directed mutagenesis. This is the first expression of glycinin-like storage protein in
E. coli.
Fifty-five patients with antibodies to HCV and chronic liver disease have been enrolled in the study. Thirty-four patients were treated with recombinant alpha interferon (IFN, 3 MU daily for 10 days ...followed by 3 MU twice/week for 3 months), and were compared to 21 untreated controls. Alanine aminotransferase (ALT) normalization was observed in a significant proportion of treated patients (52.9%), but 66.6% of them experienced a relapse after discontinuation of the therapy. The evaluation of the early ALT behavior after the 10 days priming with daily IFN administration was useful in predicting the response. The administration of a second IFN course with the same schedule and duration as the first course did not increase the efficacy of the treatment. Increased dosage and/or prolonged administration are probably required.
The aim of this study was to investigate the effect of periodontal associated bacteria in plaques adhering to peri-implant tissue of osseointegrated implants. 24 implant sites in 21 subjects placed ...with IMZ® implants, were selected for this study. Plaques adhering to implants were examined by immunoslot blot and immunoblot assay with monoclonal antibodies that specifically recognized Porphyromonas gingivalis, Prevotella intermedia serotypes I, II, Prevotella melaninogenica and Actinobacillus actinomycetemcomitans serotype b. We also investigated the relationship between the organisms and clinical parameters. According to the result of this examination, periodontal associated bacteria were detected in 11 patients and 13 patients had undetectable. Correlation between the detected periodnotal associated bacteria and clincal parameters in the positive group were observed in probing depth and bleeding on probing.
One hundred and four clinical specimens from provincial public health laboratories were tested for antibody to hepatitis C virus (HCV) envelope protein (anti-E2). To evaluate the effect of ...hypervariability of E2 region on anti-E2 assay, 49 recombinant immunoblot assay (RIBA) 3.0 positive samples were genotyped. All 49 genotyped samples were positive for anti-E2. Eight of 12 (67%) indeterminate, HCV RNA positive samples were anti-E2 reactive. Nine of 30 (30%) indeterminate, HCV RNA negative samples were also positive for anti-E2. Anti-E2 was detected in two of 13 (15%) RIBA-negative and enzyme immunoassays-positive samples. Although small number of samples were tested, the results showed that it may be possible to resolve indeterminate samples with the anti-E2 assay.
An immunoblot (Western) assay was developed employing a species-specific monoclonal antibody to a 43 kDa Mycoplasma pneumoniae membrane polypeptide and a species-specific monoclonal antibody to 29 ...kDa Legionella pneumophila outer membrane protein. This assay could simultaneously detect these two different antigens directly in sputum. The 43 kDa M. pneumoniae antigen was detected by this assay in each of three M. pneumoniae culture-confirmed sputum specimens. In addition, the 29 kDa L. pneumophila antigen was detected in three of three L. pneumophila culture-confirmed sputum specimens. Neither of these two specific antigens were detected in induced sputum specimens from ten normal individuals.
Anti-hepatitis C virus antibody screening of blood donors in different countries revealed prevalences ranging from 0.4-1.4%. These results were obtained with an enzyme immunoassay based on a ...recombinant hepatitis C virus antigen. We applied a specific inhibition assay (neutralization assay) and a recombinant immunoblot assay to determine the specificity of positive reactions in the enzyme immunoassay. Of 2836 blood donor sera tested, 10 (0.35%) were reactive in the enzyme immunoassay, however, only 3 sera (0.1%) proved to be specifically anti-HCV positive in the inhibition assay. The recombinant immunoblot assay gave similar results. The prevalence of anti-hepatitis C virus antibodies among blood donors has been overestimated in recent publications. Furthermore the high rate of false positives in the enzyme immunoassay may explain reports claiming that only a minor part of EIA positive blood units transmitted the hepatitis C virus to recipients. The inhibition assay was also applied to sera of haemophiliacs and of patients with hepatopathy which had reacted positively in the anti-hepatitis C virus antibody enzyme immunoassay. The anti-hepatitis C virus specificity was confirmed for all sera from the haemophiliacs group (100%) and for 77% of the hepatopathy patients group. Thus, the anti-hepatitis C virus enzyme immunoassay has a high predictive value when it is used to screen groups with high risks of parenteral hepatitis C virus infection, however, its predictive value is very low when it is used for blood donor screening.
ELISA, the usual screening test for HCV infection, may yield nonspecific results. Most laboratories, therefore, for corroboration perform PCR. However, a negative HCV-PCR does not prove nonspecifity ...of the initial ELISA test and therefore an immunoblot has to be performed. RIBA-II was used for that purpose for several years, but suffers from a high number of indeterminate results. We therefore compared RIBA II results of 75 sera with those of RIBA III, Matrix, Western Blot (Murex) and INNO-LIA tests. Of 34 sera that were positive in RIBA II, all were also positive in the four other immunoblots. Similarly, these 4 tests showed concordantly positive results in 13 of 27 RIBA II indeterminate sera. In the remaining 14 (RIBA II indeterminate) sera the four immunoblots displayed no uniform results but various combinations of positive, indeterminate and even negative results. Similar results were found with 13 RIBA II negative (ELISA positive) sera. These data may indicate less sensitivity as well as some nonspecificity of some of the immunoblots. In general, however, the four newly developed immunoblots proved to be more sensitive than RIBA II. This obviously is not caused by the inclusion of the NS
5-antigen but by improvement of the conventional antigens. Only one serum was found in which the NS
5-band was crucial for its positivity. In conclusion, for corroboration of some HCV-ELISA positive, PCR negative sera, more than one immunoblot may be necessary.
•Myositis-specific (MSA) and -associated (MAA) antibodies in systemic sclerosis (SSc).•The prevalence of MSA/MAA, MSA and MAA were 17%, 8.0% and 9.7%, respectively.•The isolated prevalence of each ...antibody was low (inferior to 5%).•MAA positivity was associated with ILD and myositis.•No clinical associations were found with MSA positivity.
In systemic sclerosis (SSc) and idiopathic inflammatory myopathies (IIM), auto-antibodies are used in daily practice as potent biomarkers of clinical phenotypes. This study aimed at estimating the prevalence of myositis-specific (MSA) and myositis-associated (MAA) auto-antibodies in a well-characterised SSc patients cohort using two different immunoblot assays, and studying their clinical associations.
In this cross-sectional study, the sera of 300 consecutive patients were tested at the same time with myositis antibodies Euroimmun® and D-tek® immunoblot assays.
Prevalence of MSA/MAA, MSA and MAA were 17.0%, 8.0% and 9.7%, respectively. When combining results of both tests, anti-PM/Scl 100 were found in 5.0% (95% confidence interval 2.8; 8.1); anti-PM/Scl 75 and anti-TIF1γ in 3.7% (1.8; 6.5); anti-Ku 3.0% (1.4; 5.6); anti-MDA5 in 1.3% (0.4; 3.4); anti-Mi-2 β, anti-NXP2, anti-PL-7 and anti-SRP in 0.7% (0.08; 2.4); anti-EJ and anti-PL-12 in 0.3% (0.01; 1.8) of patients. No reactivity against SAE1, Jo-1 or OJ was observed. Anti-PM/Scl 75 antibodies were associated with interstitial lung disease (80% vs. 42%) and myositis (27% vs. 3%); anti-Ku antibodies were associated with myositis (33% vs. 3%).
In this cross-sectional study of 300 SSc patients, the prevalence of MSA/MAA, MSA and MAA using immunoblot assays were 17.0%, 8.0% and 9.7%, respectively. MAA positivity was associated with ILD and myositis, but this study did not highlight any clinical associations with MSA positivity.
PDF
