Diagnosis of viruses infecting essential-oil plant crops by using immunoenzyme analysis

The use of the EIA point technique permitted the detection of viruses affecting essential-oil plants. The method is simple, highly specific, sensitive and can be used for checking seedlings prior to planting.     

Development of a diagnostic test system for detecting soluble staphylococcal antigens by an immunoenzyme method

ELISA is used for detecting the soluble staphylococcal antigen in patients with purulent septic infections. The optimum conditions for the assay have been established: the dose of staphylococcal gamma globulin for plate sensitization should be 5.0-10.0 micrograms/ml, the pH of the buffer solution 9.6-10.0, the time and temperature of incubation 18-20 hours at 4 degrees C or 5 hours at 37 degrees C. The possibility of using plates manufactured in the USSR has been shown. The sensitivity of the above diagnostic test system is 0.005 microgram/ml.     

Determination of ovalbumin-specific IgG antibodies in mouse sera by immunoenzyme analysis using horseradish peroxidase-conjugated staphylococcal protein A

The article deals with the possibility of using staphylococcal protein A conjugated with horse-radish peroxidase for the detection of specific IgG-antibodies to ovalbumin in mice by the indirect and competitive EIA techniques. Studies on specifying the parameters of the EIA system for the detection of specific IgG-antibodies are in progress.     

An immunoenzyme test system for determining the staphylococcal exotoxin of toxic shock

A highly sensitive and specific enzyme immunoassay system for the determination of staphylococcal toxic shock exotoxin (TSE), permitting the detection of TSE at a concentration of 5-10 ng/ml, has been developed. The possibility of using this assay system for the selection of TSE-producing strains has been shown. 84% of staphylococcal strains under study have been found to produce TSE.     

Approaches to determining the extent of tetanus immunity using an immunoenzyme method

The immunological survey of 3435 cattle-breeders of the Rostov region was carried out with the use of the enzyme immunoassay (EIA). The survey made it possible not only to establish the intensity of collective anti-tetanus immunity, but also to evaluate the quality of immunization. Among subjects with the known history of immunization the protective antitoxic titer was detected in 96.8 +/- 1.2% of cases and among subjects whose immunization history was unknown, in 75.3 +/- 0.8% of cases.     

Choice of the method for isolating conjugates of staphylococcal protein A with peroxidase for immunoenzyme analysis

An attempt to obtain the preparations of peroxidase-labeled staphylococcal protein A, intended for use in the enzyme immunoassay, by the glutaraldehyde method has failed. The modified periodate method permitting the preparation of active conjugates of staphylococcal protein A with peroxidase has been developed.     

Detecting staphylococcal enterotoxin B using an automated fiber optic biosensor

The Man-portable Analyte Identification System (MANTIS), the first fully automated, self-contained, portable fiber optic biosensor, was utilized for the detection of Staphylococcal Enterotoxin B (SEB), a bacterial toxin produced by Staphylococcus aureus that commonly causes food poisoning. Because of its remarkable toxicity and stability, SEB is considered a prime threat as a biological weapon of mass destruction. The assay for SEB was used to evaluate the MANTIS' ability to function in the presence of various environmental interferents. The sensor could reliably detect SEB spiked into liquid samples containing a variety of smoke particles. However, substantial interference occurred when SEB was mixed into matrices capable of adsorbing SEB, such as 1% solutions of clay, topsoil, or pollen. Of equal importance, none of the interferents produced false positives in the MANTIS. The MANTIS demonstrated the capability to perform simultaneous immunoassays rapidly in the field with little or no user intervention.     

Diagnosis of haemophilia B using the polymerase chain reaction

The polymerase chain reaction (PCR) was used to amplify specific DNA sequences within the factor IX gene of haemophilia B patients and their relatives. Three of the amplified fragments contain polymorphic sites, which can be used as markers in segregation analyses. These restriction fragment length polymorphisms (RFLPs) were until recently detected by Southern blotting after digestion with the restriction enzymes Taq I, Dde I and Xmn I. All three RFLP's are located in introns of the factor IX gene and together are informative in approximately 70% of all cases. Each of the polymorphisms was successfully used in carrier detection studies after amplification of the relevant fragments. This method is also suitable for rapid antenatal diagnosis. Additionally we were able to amplify all eight exons of the factor IX gene including the splice junctions and a part of the 5'-region. Large deletions or insertions can be detected without further analysis. Several possibilities for the rapid detection of point mutations after DNA amplification have been described recently. The complete amplification of all functional parts of the Factor IX gene in combination with these new techniques should enable us to detect the majority of mutations leading to haemophilia B.     

The standardization of conjugates for the indirect solid-phase immunoenzyme analysis reaction

The comparative study of several batches of conjugates has revealed that enzyme immunoassay techniques can be used for the standardization of conjugates by their affinity level. The study has shown that this can be done only if the concentration of specific antibodies in the conjugate is known and the amount of the conjugate is in excess to that of the antigen adsorbed on the plate.     

Possible diagnosis of Pseudomonas aeruginosa infection by an immunoenzyme method using pyoimmunogen as the antigen

The possibility of detecting P. aeruginosa antibodies in patients by means of indirect solid-phase EIA techniques is shown. This assay is carried out with the use of reagents produced in the USSR: polystyrene assay plates manufactured by the Lenigrad Medpolymer Works are used as carriers, P. aeruginosa vaccine (pyoimmunogen) obtained under semi-industrial conditions at the Mechnikov Central Research Institute for Vaccines and Sera is used as antigenic complex and the commercial preparation produced by the Gamaleia Research Institute of Epidemiology and Microbiology serves as conjugate. The studies have revealed that in 95% of cases the level of antibodies in the sera of patients with acute destructive pneumonia accompanied by pleural empyema, abscesses of internal organs and acute hematogenic osteomyelitis is essentially higher than the level of "normal" antibodies in healthy donors from whom biologically confirmed P. aeruginosa cultures can be isolated. In the groups of patients with similar nosological forms of diseases caused by other infective agents such difference in antibody titers is not detected. These results suggest that the detection of antibodies to P. aeruginosa in patients' sera by means of EIA can be used as an additional test for the diagnosis of P. aeruginosa infections.     

Serological diagnosis of suppurative-septic diseases of staphylococcal etiology

The parallel examination of osteomyelitis patients by means of the passive hemagglutination (PHA) test with the use of antigenic erythrocyte diagnostic agents, prepared on the basis of hydrochloric acid extract from Staphylococcus aureus strain 209P and teichoic acid extract from S. aureus strain Wood 46 has revealed that these diagnostic agents are practically equal in their diagnostic effectiveness. In the examination of endocarditis patients measurement of the total antibody activity in the PHA test with diagnosticum on the basis of strain 209P has proved to be a more sensitive method, whereas in osteomyelitis patients the total IgG activity has been more accurately measured by ELISA. The treatment of sera with 2-mercaptoethanol essentially decreased the diagnostic effectiveness of the PHA test. The study has shown the diagnostic value of not only IgG but also of IgM antibody measurements.     

Study of the interaction of low-density lipoproteins with immobilized fibrinogen and fibronectin using an immunoenzyme assay

The interaction of serum low density lipoproteins (LDL) with fibrinogen, fibronectin and fibrinogen-fibronectin complex immobilized on polystyrene was studied. LDL binding by these proteins was assessed by enzyme-linked immunosorbent assay. It was shown that LDL interacts with both immobilized fibronectin and immobilized fibrinogen. The fibrinogen-fibronectin complex bound a greater amount of LDL than either component alone, in the same concentration as in the complex. The binding of LDL with immobilized fibronectin or fibrinogen increased in the presence of low concentrations of diluted fibrinogen or fibronectin, respectively, which suggests the formation of the three-component complex on the surface. The formation of fibrinogen-fibronectin-LDL complexes during homeostasis may promote the accumulation of LDL in the vascular wall.     

Evaluation of postvaccinal pertussis immunity by using immunoenzyme analysis

The effectiveness of adsorbed DPT vaccine manufactured in the USSR, evaluated by its capacity of inducing the formation of the main classes of immunoglobulins and by the duration of immune response to the acellular complex of protective antigens (pertussis toxin and agglutinogen-2), was studied with the use of modified EIA. Out of 273 children immunized with adsorbed DPT vaccine in the course of this study, 87.2% had IgG-antibodies, 14.1% had IgA-antibodies and 3.2% of the children had IgM-antibodies. The level of immunity in children having received the full course of immunization with adsorbed DPT vaccine was significantly higher in comparison with children given only the primary course of immunization and nonimmunized children of the same age. Antipertussis immunity was found to decrease two years after the completion of the course of immunization with adsorbed DPT vaccine and in children over 5-6 years of age. Adsorbed DPT vaccine prevented the disease, but not infection. The level of postinfection immunity was higher than that of postvaccinal immunity.     

Correlation of titers of antibodies to the measles virus by an immunoenzyme method and in the hemagglutination inhibition reaction

The authors analyze the results of comparative studies on 15 paired sera from children with suspected measles, of 32 sera from children and adolescents aged 1.5 to 16 immunized against measles, and of 21 sera from adults aged 19 to 86 with a history of the disease. EIA proved to be more sensitive than HAIT: the detection rate of positive sera was higher, as were the titers of antibodies detected by it, in examinations of the sera from vaccinated children and the adults. Analysis of the distribution of sera with different titers of antibody to measles virus in EIA and HAIT has revealed a correlation between the titers in the sera with high antibody levels. In the cases with low antihemagglutinin titers, no correlation between the titers determined in the two tests has been observed.     

Development of an immunoenzyme method for detecting Francisella tularensis

The conditions permitting the determination of F. tularesis cells by means of the enzyme immunoassay (EIA) in 3-5 hours have been established. Ways for enhancing the reliability of results obtained in the assay of the least possible amount of the test material have been proposed. The sensitivity and specificity of the rapid EIA technique permitting the determination of F. tularensis cells at a concentration of 20 000 cells/ml in the presence of other bacterial cells in 100-fold excess have been shown.