An immunoenzyme test system for determining gaprin in the air]

A highly specific and sensitive enzyme immunoassay system for the quantitative determination of haprin in the air at a concentration of not less than 0.0001 mg of haprin per m3 of air has been developed. The assay system has been approved at the Svetloiarsk plant for the production of vitamin protein concentrate and can be used for the control and evaluation of haprin pollution of the environment at areas adjoining microbiological plants.     

Methods of enhancing the specificity of immunoenzyme test systems for the determination of human IgE

The sensitivity and specificity of an enzyme immunoassay (EIA) system for the determination of total IgE in humans increases if polystyrene plates are sensitized with swine gamma-globulin and the conjugate is prepared with the use of sheep gamma-globulin obtained from the commercial preparation of sheep antiserum to human IgE. Such system becomes even more specific and sensitive if sheep antibodies to human IgE, purified by affinity chromatography, are used both for the sensitization of polystyrene plates and for the preparation of the conjugate. This conjugate is necessary for the development of EIA systems intended for the determination of specific human IgE-antibodies to various allergens.     

Quantitative evaluation of serum IgG subclass levels by an immunoenzyme method

We developed a enzyme linked immunosorbent assay (ELISA) for measuring IgG subclasses concentration in serum. For this we used monoclonal antibodies. The specificity of these antibodies was evaluated with a panel of myeloma proteins belonging to the 4 IgG subclasses. The ELISA was sensitive (allowing the detection of subclasses at ng level) and accurate (inter-assay coefficient of variation of 14%). Using the WHO serum 67/97 as reference, we determined the concentration of IgG subclasses in a pool of sera. In addition concentrations were measured in 69 healthy adults to study the distribution of each IgG subclass. A good correlation (r = 0.78) was obtained between the sum of the subclasses measured by ELISA and total IgG measured by immunonephelometry.     

An immunoenzyme test system for the detection of antibodies to Campylobacter]

An enzyme immunoassay system for the detection of antibodies to bacteria of the genus Campylobacter in human blood serum has been developed. The system is based on the use of ethanol-treated C. jejuni and C. coli whole cells as antigen. The study of sera obtained from healthy donors in this assay has made it possible to establish the value of the tentative diagnostic titer: 320.     

Immunochemical study of the heterogeneity of trophoblastic beta 1-glycoprotein

It has been found that in mild acid (pH 6.6) and mild-alkaline media (pH 7.7) both pregnancy proteins form complete precipitates. In more alkaline buffer solutions the form of alpha 2-glycoprotein (alpha 2-GP) precipitate is preserved, while trophoblastic beta 1-glycoprotein (TBG) shows three immunochemically identical components with different electrophoretic mobility. The form with beta-globulins mobility predominates, and minor fragments are presented by alpha- and gamma-components. All TBG forms are clearly seen at pH 8.6. In more alkaline medium (pH 10.0) the clarity of the precipitates drastically decreases. It is shown that heparin introduction into the gel of first dimension electrophoresis increases anode electrophoretic mobility of both proteins at polysaccharide concentration of at least 0.1 mg/ml. Large amounts of heparin cause the increase in TBG alpha-component precipitate area and the decrease in the form with beta-globulins mobility. At the same time alpha 2-GP precipitate area and form remain unchanged.     

Characteristics of the carbohydrate component of trophoblastic beta1-glycoprotein

By use of the methylation method, trophoblast-specific beta 1-glycoprotein (TSG, SP-1) has been shown to contain N-glycosidically linked oligosaccharide chains of the N-acetyl lactosamine type. Methylation analysis of TSG fractions which have various affinity to concanavalin A indicates that the diantennary carbohydrate chains prevail in the glycoprotein molecule.     

Immunoautoradiographic determination of beta-1 gamma globulin in the blood serum of patients with trophoblastic tumors]

The authors determined beta1-G-globulin (beta1GG) in the sera of patients with different malignant tumours and of normal donors by means of the immunodiffusion method (ID) and immunoautoradiography (IAR). In the ID-negative sera beta1GG was revealed by means of IAR in 7 out of 8 patients with chorionepithelioma of the uterus and in 1 out of 7 patients with teratomblastoma of the testis before the treatment. After the treatment the beta1GG was determined in 7 out of 21 patients with chorionepithelioma of the uterus. At the early stage of trophoblastic tumours of the uterus beta1GG could be found in 77.7% of cases by means of IAR and in 16.8% of cases by means of the ID method.     

The comparative evaluation of spectrophotometric and chemiluminescent detection in an immunoenzyme test of antibodies to the antigens of the bovine leukosis virus]

Comparative evaluation of the sensitivity limit in the detection of antibodies to bovine leukemia virus in the enzyme immunoassay with the use of chemiluminescent and spectrophotometric detection techniques was carried out. In this assay 3-amino-1,4-phthalazinedion was used as chemiluminescent substrate and ortho-phenylenediamine, as chromogenic substrate. The chemiluminescent signal was registered by means of a special luminometer designed at the Institute of Biochemistry (Lithuanian Acad. Sci.). The use of the chemiluminescent substrate permitted the detection of proteins in amounts 2-3 times lower than those detected by the spectrophotometric technique.     

Quantitative evaluation of immunohistochemical staining in gastrointestinal stromal tumors

OBJECTIVE: To evaluate the differences among pathologists' interpretations in gastrointestinal stromal tumors (GISTs) and to demonstrate the usefulness of quantitative pathology in the assessment of immunohistochemical staining. STUDY DESIGN: Twenty GISTs were separately evaluated by 4 pathologists by the visual estimation method using a 6-antibody panel. Each case was then quantitatively measured with a computer-assisted image analysis system by 2 pathologists. Cohen's kappa test was performed for statistical analysis. RESULTS: All GISTs showed some degree of expression of CD 117, CD34, SMA and Ki-67. No case was immunoreactive for desmin or S-100 protein. There were remarkable differences in the pathologists' visual estimations. Moreover, the discrepancies between visual and quantitative methods were noteworthy. The differences in interpretations showed the greatest variability for Ki-67, which is known to be related to poor prognosis. CONCLUSION: Quantitative pathology in assessment of immunohistochemical staining of GISTs may improve the consistency in the interpretation of staining results and provide some degree of reproducibility.     

Informativeness of angiography in trophoblastic tumors of the uterus

A study was made of the resolving power of general and selective pelvic angiography with regard to specified diagnosis of trophoblastic tumors of the uterus. The informative value of general pelvic angiography was estimated by the results of angiomorphological correlations in 81 operated patients. The significance of selective pelvic angiography performed in 152 women was based on comparison of its results with those of pharmacoangiography supplemented by hysterography in 86 patients. The correctness of diagnosis in 26 of them was confirmed by operative findings. Analysis of the investigations has shown that general pelvic angiography allowed the detection of a tumor, its size and site in the uterus in 48% of the patients only. Selective pelvic angiography combined with pharmacoangiography was shown to be a method extending the diagnostic potentialities of pelvic angiography. In the authors' experience, it permitted obtaining a direct contrast image of a trophoblastic tumor or excluding its presence in the uterus of 85% of the examinees.     

Detection of trophoblastic beta 1-glycoprotein in tumor tissue and blood in ovarian neoplasms

Using immunochemical analysis methods (the reaction of precipitation in agar, immunoenzymatic method, immunofluorescence), trophoblastic beta 1-glycoprotein (TBG) concentration in tumour tissue and in the blood serum of patients with ovarian cancer was studied. By rabbits immunization with glycoprotein fraction of ovarian adenocarcinoma, dissolved in 0.6 M sulfosalicylic acid, the authors obtained antibodies to TBG. Immunoenzymatic method showed, that TBG level is raised during ovarian cancer (more than 3 micrograms/l): in 18% of tumour extracts, in 12.5% of blood sera samples and in 41.6% of cases in ascites fluid. Utilizing indirect immunofluorescence method morphological structures of trophoblastic type were identified in paraffin sections of ovarian adenocarcinoma. The authors suppose, that such structures may be responsible for TBG biosynthesis in ovarian tumours.     

Diagnostic test systems based on the immunoenzyme method in leprosy

Diagnostic test systems for the detection of IgG and IgM to Mycobacterium leprae in the blood sera of leprosy patients and armadillos experimentally infected with M. leprae have been developed on the basis of the indirect immunoperoxidase assay. The possibility has been shown of prognosing the activity of the leprotic process in leprosy patients and the results of the experimental infection of armadillos by the dynamic increase of antibody reactions with the development of the infection.     

Substrate specificity of the protease that processes human interleukin-1 beta

The substrate specificity of the protease which generates mature human interleukin-1 beta (IL-1 beta) from pro-interleukin-1 beta was investigated using synthetic peptide substrates and recombinant pro-IL-1 beta. The requirement of an L-aspartate in the P-1 position was confirmed together with the need for a small hydrophobic residue in the P-1' position (Gly or Ala). It was shown that the enzyme can tolerate conservative substitutions in the P-2 and P-2' positions. We found little difference in the enzyme's ability to cleave denatured and native pro-IL-1 beta, indicating that tertiary structure recognition is not involved in binding. The enzyme did, however, require a peptide of more than six amino acids for cleavage to occur. These results conclusively demonstrate the unusual specificity of this protease.     

The DNA content in the cell nuclei of trophoblastic tumors

The DNA content in cytotrophoblast (CTB) and syncytiotrophoblast (STB) cell nuclei was assayed in tissue sections of 7 hydatidiform moles (HM) and 27 choriocarcinomas (CH). The procedure involved Feulgen's reaction and scanning cytophotometry. The analysis of summarized histograms showed the DNA distribution in CTB cell nuclei, on the one hand, and that in STB, on the other, to differ significantly in both the tumors. The HM studied cases were referred to as two subtypes on the basis of such parameters as modal class value, its ploidy and degree of nuclear poly- and heteroploidy of CTB and STB. These characteristics were used to identify three patterns of CH. A pronounced modal class (2c--4c) was typical of type 1. A wider range of modal class (2c--10c or 4c--8c) was observed in type 2. Type 3 of tumor was characterized by a pronounced polyploidy with the absence of the modal class. The analysis of individual CTB and histograms showed no significant differences between HM and CH with respect to the DNA content. An increase in the share of highly polyploid cells was associated with a shorter survival of patients.     

Characterization of the interleukin-1beta system during porcine trophoblastic elongation and early placental attachment

Conceptus-uterine communication is established during trophoblastic elongation when the conceptus synthesizes and releases estrogen, the maternal recognition signal in the pig. Interleukin-1beta (IL-1beta) is a differentially expressed gene during rapid trophoblastic elongation in the pig. The current investigation determined conceptus and endometrial changes in gene expression for IL-1beta, IL-1 receptor antagonist (IL-1Rant), IL-1 receptor type 1 (IL-1RT1), and IL-1 receptor accessory protein (IL-1RAP) in developing peri- and postimplantation conceptuses as well as uterine endometrium collected from cyclic and pregnant gilts. Conceptus IL-1beta gene expression was enhanced during the period of rapid trophoblastic elongation compared with earlier spherical conceptuses, followed by a dramatic decrease in elongated Day 15 conceptuses. IL-1RT1 and IL-1RAP gene expression was greater in Day 12 and 15 filamentous conceptuses compared with earlier morphologies while IL-1Rant gene expression was unchanged by conceptus development. The uterine lumenal content of IL-1beta increased during the process of trophoblastic elongation on Day 12. Uterine IL-1beta content declined on Day 15, reaching a nadir by Day 18 of pregnancy. IL-1beta gene expression in porcine conceptuses was temporally associated with an increase in endometrial IL-1RT1 and IL-1RAP gene expression in pregnant gilts. Endometrial IL-1beta and IL-1Rant gene expression were lowest during Days 10-15 of the estrous cycle and pregnancy. The temporal expression of IL-1beta during conceptus development and the initiation of conceptus-uterine communication suggests conceptus IL-1beta synthesis plays an important role in porcine conceptus elongation and the establishment of pregnancy in the pig.