LabVIEW-based control and acquisition system for the dosimetric characterization of a silicon strip detector.

The aim of this work is to present a new data acquisition, control, and analysis software system written in LabVIEW. This system has been designed to obtain the dosimetry of a silicon strip detector in polyethylene. It allows the full automation of the experiments and data analysis required for the dosimetric characterization of silicon detectors. It becomes a useful tool that can be applied in the daily routine check of a beam accelerator.

Characterization of the Aspergillus nidulans aspnd1 gene demonstrates that the ASPND1 antigen, which it encodes, and several Aspergillus fumigatus immunodominant antigens belong to the same family.

For the first time, an immunodominant Aspergillus nidulans antigen (ASPND1) consistently reactive with serum samples from aspergilloma patients has been purified and characterized, and its coding gene (aspnd1) has been cloned and sequenced. ASPND1 is a glycoprotein with four N-glycosidically-bound sugar chains (around 2.1 kDa each) which are not necessary for reactivity with immune human sera. The polypeptide part is synthesized as a 277-amino-acid precursor of 30.6 kDa that after cleavage of a putative signal peptide of 16 amino acids, affords a mature protein of 261 amino acids with a molecular mass of 29 kDa and a pI of 4.24 (as deduced from the sequence). The ASPND1 protein is 53.1% identical to the AspfII allergen from Aspergillus fumigatus and 48% identical to an unpublished Candida albicans antigen. All of the cysteine residues and most of the glycosylation sites are perfectly conserved in the three proteins, suggesting a similar but yet unknown function. Analysis of the primary structure of the ASPND1 coding gene (aspnd1) has allowed the establishment of a clear relationship between several previously reported A. fumigatus and A. nidulans immunodominant antigens.

Immunoblotting patterns in the serodiagnosis of aspergilloma: antibody response to the 90kDa Aspergillus fumigatus antigen.

At present there are no accepted criteria to assess the usefulness of Western blot assays for the serodiagnosis of aspergilloma. An Aspergillus fumigatus cytosolic fraction complex (CFC) composed of four proteins (p90, p60, p40, and p37) has been identified. The usefulness of Western blotting with CFC antigens for the serodiagnosis of aspergilloma was evaluated in 25 patients with well-established diagnoses and in 94 controls. The most consistently reactive antigen was p90 (92% of patients with aspergilloma), followed by p40 (76%) and the entire CFC taken together (76%). With these data, interpretive criteria for positive and negative immunoblots were established, with p90 indicated as a helpful marker of aspergilloma.

An immunodominant 90-kilodalton Aspergillus fumigatus antigen is the subunit of a catalase.

We have identified, purified, and characterized structurally and functionally a 90-kDa immunodominant antigen associated with the water-soluble fraction of Aspergillus fumigatus. This antigen is recognized by 90.3% of serum samples from patients with aspergilloma and should be considered either by itself or better in combination with other purified antigens as a candidate for developing a standardized immunoassay for the detection of aspergilloma. p90 is a glycoprotein containing at least two two N-linked sugar chains of 2 and 5 kDa, respectively, which are not necessary for its reactivity with aspergilloma serum samples. Using specific anti-p90 rabbit serum, we have demonstrated that under native conditions, p90 exists in oligomeric form and has associated catalase activity. This activity is resistant to extreme temperatures (> 60 degrees C), reducing agents (40 mM dithiothreitol), high concentrations of denaturing agents such as 8 M urea and 8% sodium dodecyl sulfate, and treatments with ethanol-chloroform-water (5:3:10 [vol/vol]) mixtures.

Aspergillus fumigatus antigens.

Cytosolic fractions of mycelial extracts from Aspergillus nidulans, A. flavus, and three different isolates of A. fumigatus, grown to stationary phase in Czapek-Dox-AOAC medium, were tested by immunoblotting for the presence of antigens reactive to 80 serum samples from aspergilloma patients. Fifty control serum samples were used to determine the specificity of the reactions. In the A. fumigatus cytosolic fraction a group of four main antigenic bands (p90, p60, p40 and p37) was consistently recognized (in total or partial form) by 90% of the serum samples from the aspergilloma patients. This group of antigens was designated as the 'cytosolic fraction complex' (CFC). As confirmed by two-dimensional electrophoresis followed by immunoblotting with aspergilloma serum samples, each of the four antigenic bands is formed of several isoforms of acidic glycopeptides with slightly different pls. All the isoforms are at least N-glycosylated, as demonstrated by endoglycosidase H removal of a considerable amount of sugar residues. The relationship of these antigens with certain other A. fumigatus antigens previously reported in the literature, and their potential use in the immunodiagnosis of aspergilloma, are discussed.

Variability of Aspergillus nidulans antigens with media and time and temperature of growth.

The influence of culture medium and time and temperature of growth on the appearance of Aspergillus nidulans antigens was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by silver staining or Western blot (immunoblot), of the proteins present in total cellular extracts or culture supernatants. Samples in the exponential, deceleration, and stationary growth phases were selected by biochemical, morphological, and ultrastructural criteria. Protein and antigen patterns (detected with rabbit antibodies) from total extracts were very similar in all cases, and the major differences observed seemed to depend on the age of the cultures. Culture supernatant patterns were highly dependent on the type of medium (complex or defined) and the age of the culture. Temperature did not significantly influence these results. The reproducible reactivity of selected human sera from aspergilloma-affected individuals was strictly associated with the use of defined media, especially Czapek Dox-AOAC, in both total extracts and culture supernatants. Extended growth times were necessary in the case of metabolic antigens (those obtained from culture supernatants). Screening of a battery of 10 selected human serum samples from patients with aspergilloma or invasive aspergillosis demonstrated that two of the antigens (96 to 98 and 45 kDa) from stationary-phase culture supernatants in Czapek Dox-AOAC medium were consistently reactive. When considered together as one unit, both antigens reacted with more than 50% of the sera, and at least one or the other of the antigens reacted with more than 90% of the sera. Less consistent results were obtained for two somatic antigens (from total cell extracts) of 45 to 50 and 20 to 22 kDa.

Immunogenic potential of Aspergillus nidulans subcellular fractions and their polypeptide components.

Cell-free extracts of the ascomycetous fungus Aspergillus nidulans were separated into three subcellular fractions: cell walls, total membranes and cytosol, and two different immunization protocols were used to raise antibodies against them in 12 New Zealand rabbits. The immune response was followed over time by dot and Western blot analyses to determine the immunogenic potential of each individual fraction and their polypeptide components. The IgG fractions, purified from pools of the best sera, were used to analyze in detail the antigenic composition of A. nidulans mycelium. The fast immunization protocol provided a much earlier response and higher sera titres. Cytosols and membranes were more immunogenic than cell walls and, in most cases, a positive correlation was shown between the titre of each serum and the number of detected antigens. The polypeptides of A. nidulans included six major immunodominant antigens of the molecular weights ranging between 13 and 200 kDa.

Analysis of Aspergillus nidulans conidial antigens and their prevalence in other Aspergillus species.

Aspergillus nidulans is an ascomycetous fungus that reproduces asexually by forming multicellular conidiophores and uninucleate spores called conidia. These elements constitute the main vehicle for the transmission of this and other pathogenic Aspergillus species and are the starting point of the different forms of aspergillosis. In order to use A. nidulans as a potential source of useful antigens for the immunodiagnosis of these diseases, we have examined the total protein composition of conidial extracts of this fungus by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis in gels of different percent T. Injection of SDS-extracted conidial proteins into rabbits allowed us to raise a battery of polyclonal antibodies which have defined some important immunogenic polypeptides. Several of these immunogens were both present in mycelial extracts and recognized by antimycelium antibodies. Four of them, designated cdA, cdB, cdC, and cdE, were also found in conidial extracts of other pathogenic Aspergillus species. Only cdE was undetectable in cell extracts of the nonrelated species Fusarium culmorum and Phycomyces blakesleeanus.