Agar Gel Double Diffusion Research Articles

The value of serum precipitins against specific antigens in patients diagnosed with hypersensitivity pneumonitis - retrospective study.

IntroductionHypersensitivity pneumonitis (HP) is the third most common interstitial lung disease, and is often under-recognized, especially in patients who are not aware of their occupational or environmental contact with organic antigens. The aim of the present study was to assess the results of serum specific IgG antibodies (ssIgG) in HP patients and their correlation with clinical data.Material and methods128 HP patients, median age 53 years, participated in the study. The control group consisted of 102 patients with interstitial lung diseases (ILDs) other than HP. Assessment of pretreatment ssIgG to thermophilic actinomycetes and protein antigens from bird droppings (pigeons, hens, ducks, parrots, turkeys) was performed by double diffusion in agar gel according to Ouchterlony method.ResultsPositive precipitins were obtained in 57% of all HP patients and in 61% of those exposed to above mentioned antigens. Positive results in the control group were obtained in 7% of patients. Sensitivity of ssIgG in HP group was 0.57 and specificity 0.93. Precipitins to at least one bird antigen was confirmed in 64% of HP patients exposed to birds. Precipitins to thermophilic bacteria were found in 29% of HP patients exposed to hay or hay products.ConclusionsThe results of the study indicate that ssIgG against birds’ allergens were the valuable diagnostic tool in HP patients. Low-rate of confirmation of ssIgG to thermophilic bacteria in patients exposed to hay or hay products indicate that other microorganisms, most likely molds, could be responsible for the disease development.

Intra-abdominal Aspergillosis due to Aspergillus fumigatus in a Dog

A 3 year-old, spayed female, mixed-breed dog presented with abdominal distension due to a large mass, detected during abdominal palpation and confirmed by abdominal imaging. Cytological examination of the mass was suggestive of pyogranulomatous inflammation. During exploratory laparotomy, extensive peritoneal adhesions and multifocal nodular lesions on the liver, spleen and omentum were revealed. One week later, the dog deteriorated and was euthanized. Numerous firm masses were observed in the liver, spleen, left kidney, stomach, small and large intestine during necropsy. The lungs, heart, and ocular structures were macroscopically normal. Histopathology results (surgery and necropsy) revealed fungal hyphae enclosed in the pyogranulomatous lesions. Polymerase chain reaction (PCR) products showed 100% homology with Aspergillus fumigatus and agar gel double diffusion was positive for IgG antibodies against the same fungus.

Development of an immunochromatographic strip for detection of antibodies against duck Tembusu virus

Duck Tembusu virus (DTMUV), a novel flavivirus, causes severe disease in ducks. There is an urgent need for a rapid and effective diagnostic method to control the spread of DTMUV. We chose the envelope (E) protein from DTMUV as an antigen and combined it with colloidal gold particles as tracers to specifically detect anti-DTMUV antibodies. Based on the double-antigen sandwich format, an immunochromatographic strip (ICS) for the rapid detection of anti-DTMUV antibodies was developed. The ICS showed a high specificity and no cross-reactivity with other sera. By detecting a serially diluted duck anti-DTMUV serum, the sensitivity of the ICS was 16-fold higher than that of the agar gel double diffusion test. Moreover, the ICS was both stable and reproducible, maintaining the same performance at 4°C for at least 6 months. To evaluate the effectiveness of the ICS, 217 duck serum samples were tested with the ICS and an indirect enzyme-linked immunosorbent assay (iELISA). The consistency ratio of positive and negative results between the two methods was 97.87% and 97.06%, respectively. The agreement between the ICS and the ELISA was 97.24%. The ICS developed in this study offers a specific, sensitive, and rapid method to detect anti-DTMUV antibodies.

An immunoproteomic approach revealed antigenic proteins enhancing serodiagnosis performance of bird fancier's lung

BackgroundBird fancier's lung (BFL) caused by repeated inhalation of avian proteins is the most common form of hypersensitivity pneumonitis. However, the exact identification of proteins involved is unknown, and serological test use for diagnosis need to be standardized. The objectives of this study were (i) to identify antigenic proteins from pigeon droppings (ii) to provide information about their location in avian matrices and (iii) to produce them in recombinant proteins to evaluate their diagnostic performances. MethodAntigenic proteins of pigeon dropping extracts were investigated using 2-dimensional immunoblotting with sera from patients with BFL, asymptomatic exposed controls and healthy volunteers. We investigated the origin of these antigenic proteins by analyzing droppings, blooms and sera using a shotgun proteomic analysis. BFL-associated proteins were produced as recombinant antigens in E. coli and were assessed in ELISA with sera from patients (n=25) and subject exposed controls (n=30). These diagnostic performances were compared with those obtained by precipitin techniques (agar gel double diffusion, immunoelectrophoresis). ResultsWe identified 14 antigenic proteins mainly located in droppings and blooms. These proteins were involved in either the digestive or immune systems of pigeons.Using the recombinant BFL-associated proteins: Immunoglobulin lambda-like polypeptide-1 (IGLL1: sensitivity: 76%; specificity: 100%; AUC: 0.93) and Proproteinase E (ProE: sensitivity: 84%; specificity: 80%; AUC: 0.85), the ELISA test showed better performance than precipitin assays with pigeon dropping extracts (sensitivity: 60%; specificity: 93.3%; AUC: 0.76). ConclusionIGLL1 and ProE were identified as the biomarkers of the disease. The use of these highly standardized antigens discriminates BFL cases from exposed subjects in serological assays. The results of this study offer new possibilities for the serological diagnosis of the disease.Clinical trial registration: ClinicalTrials.gov: Identifier NCT03056404.

Aspergillusserology: Have we arrived yet?

Aspergillosis presents in various clinical forms, among them chronic pulmonary aspergillosis, which is a spectrum of disease entities including aspergilloma, chronic cavitary pulmonary aspergillosis, and chronic fibrosing pulmonary aspergillosis. Aspergillus also contributes to fungal allergy and sensitization. Analysis of the immune response to Aspergillus and its antigens is an integral part of the diagnosis of these diseases. Over the past half century, the techniques used to determine antibody titers have evolved from testing for precipitating and agglutinating antibodies by agar gel double diffusion and immunolectrophoresis to enzyme-linked immunosorbent assays using recombinant proteins as capture antigens. A resurgence of interest in the detection of immunoglobulins, primarily Aspergillus-specific IgG, has hinted at the possibility of distinguishing between colonization and invasion in immunocompromised patients with invasive aspergillosis. Even though there appears to be a greater degree of discrimination between the clinical forms of aspergillosis there is still a long way to travel. This review presents illustrative examples of where new diagnostic platforms and technologies have been applied to this intriguing spectrum of diseases.

The Australia (Hepatitis-associated) Antigen

INTRODUCTION The accidental discovery of the Australia antigen (HAA) in 1963 by Blumberg and his coworkers is a classical example of biomedical serendipity. Its recognition not only stimulated a substantial amount of research into viral hepatitis, thereby adding considerably to our knowledge about this important disease, but has had far-reaching implications in the understanding of several other hepatic and systemic diseases. Indeed, its discovery may well be seen in the future to have been the all-important breakthrough in attempts at culture of the virus as a means of vaccine preparation for future prophylactic usage. THE DISCOVERY OF THE ANTIGEN Blumberg, a cytogeneticist working in the Philadelphia Institute for Cancer Research, was conducting a systematic study of the serum of patients who had received multiple blood transfusions. Using a technique of double diffusion in agar gel (Ouchterlony method) he searched for precipitating antibodies to inherited donor serum protein antigens, particularly those to low density betalipoproteins. A precipitating antibody was identified in the serum of a haemophiliac patient which reacted with an antigen which was quite dissimilar from lipoprotein antigens. This antigen was present in the serum of an Australian aborigine, and the term Australia antigen was coined (1, 2, 3,). This antigen is almost certainly identical to Prince’s SH antigen (4), and to avoid confusion the term hepatitis-associated antigen (HAA) is now considered by many to be more appropriate (5).

Microscopic and polyclonal antibody-based detection of yellow leaf disease of arecanut (Areca catechu L.)

Phytoplasma, the pathogen of yellow leaf disease (YLD) of arecanut (Areca catechu L.) was detected by transmission and scanning electron microscopy. Tissues of YLD affected palms contained phytoplasmas in the phloem sieve elements, but not in symptomless healthy palm tissues. Phytoplasma was purified from tissues of diseased palms employing percoll density gradient centrifugation and confirmed by transmission electron microscopy. Using the purified phytoplasma preparation, a polyclonal antiserum was raised in rabbits and used for standardisation of agar gel double diffusion (Ouchterlony) test and DAC-ELISA. Clear precipitin line was observed in Ouchterlony test between the antigen from diseased palms and the pathogen-specific antibodies after 48 h incubation and only undiluted antiserum showed best result in the test. However, in ELISA, 1:10 antigen dilution and 1:400 pathogen-specific antibodies dilution produced sensitive detection of the pathogen with a difference of >3.5 times absorption values between healthy and diseased samples. The results thus confirmed the ability of antiserum to distinguish healthy and infected plants and utility of ELISA for effective diagnosis of YLD.

Utilização da serologia na identificação de <em>Ascia monuste orseis</em> (Latreille, 1819) (Lepidoptera: Pieridae)

This study aimed to obtain specific antiserum for Ascia monuste orseis (Latreille, 1819), with the aim of identifying the predators associated with this insect. The preparation of specific antiserum was performed by immunizing a rabbit via a lymph nodule injection containing antigen obtained from the maceration of 4 th and 5 th instars of A. monuste orseis caterpillars. Serological homologous tests performed using double diffusion in agar gel were positive 21 days after the first antigen inoculation. Eggs, 1 th , 2 th and 3 th instar caterpillars, pupae and adults of A. monuste orseis also reacted positively to the obtained antiserum.

Comparisons of complete RNA-2 sequences, pathological and serological features among three Japanese isolates of Arabis mosaic virus

Arabis mosaic virus lily and narcissus isolates (ArMV-Li and ArMV-Na) induced severe necrotic spots on Chenopodium quinoa, whereas ArMV butterbur isolate (ArMV-Bu) caused symptomless infection in the plant. The accumulation level of ArMV-Bu in upper non-inoculated leaves of C. quinoa was comparable to that of ArMV-Li or -Na. The agar gel double-diffusion test using an antiserum against ArMV-Li showed ArMV-Li was closely related to ArMV-Na, but not to ArMV-Bu. The RNAs-2 of ArMV-Li, -Na, and -Bu consist of 3707, 3709, and 3789 nucleotides, and they contain one open reading frame encoding a putative polyprotein of 1083, 1084, and 1122 amino acids, respectively. The overall identity of RNA-2 of ArMV-Li displayed more than 90% with ArMV-Na, but less than 70% with ArMV-Bu. A phylogenetic analysis of 2A sequences from ArMV isolates revealed ArMV-Bu was not categorized in any cluster. ArMV-Bu is a unique isolate from the point of view of pathological and serological features, and nucleotide sequence.

Anti-LKM-1 antibodies determined by use of recombinant P450 2D6 in ELISA and western blot and their association with anti-HCV and HCV-RNA.

Several subtypes of anti-liver-kidney microsome antibodies (LKM) are known. LKM-1 antibodies associated with autoimmune chronic active hepatitis recognize P450 2D6, a cytochrome P450 mono-oxygenase. The frequent association of anti-LKM-1 antibodies and hepatitis C virus (HCV) infections and the probable existence of an infectious and autoimmune form of anti-LKM-1-associated hepatitis, requiring different therapeutical strategies, necessitates the exact determination of anti-LKM-1 specificities. Therefore, we compared various antibody tests (immunofluorescence, ELISA with recombinant P450 2D6, and Western blot with recombinant and natural antigens and agargel double diffusion) with sera of 27 anti-LKM-1-positive chronic active hepatitis (CAH) patients, with 61 sera harbouring anti-mitochondrial antibodies, 100 sera each from HCV-RNA-positive and HCV-RNA-negative patients, and 50 sera of healthy persons. Western blot techniques using recombinant MS2-polymerase P450 2D6 fusion protein were found to be the most sensitive and specific method for anti-LKM-1 antibody determination in routine laboratory. The recently recognized association of anti-LKM-1 antibody and HCV infection was confirmed by the results of this study. In anti-HCV and HCV-RNA-positive patients with anti-LKM-1 antibodies there was a preponderance of males with higher mean age and lower antibody titres. The results support the hypothesis of the existence of an autoimmune as well as an infectious (HCV triggered) subgroup of anti-LKM-1-positive hepatitis.

First Report of Japanese Beetle (Popillia japonica) as a Vector of Southern bean mosaic virus and Bean pod mottle virus

The Japanese beetle, Popillia japonica (Coleoptera: Scarabaeidae), was introduced into the United States in New Jersey in 1916 and now commonly occurs in the eastern and mid-western United States. The Japanese beetle has a wide host range and feeds on more than 300 plant species within 79 families (2), but it has not been reported to be a vector of plant viruses. Southern bean mosaic virus (SBMV; genus Sobemovirus) and Bean pod mottle virus (BPMV; family Comoviridae, genus Comovirus) are transmitted by several species of leaf-feeding beetles in the families Coccinelidae and Chrysomelidae (1). Japanese beetles, collected in northwestern Arkansas with a JB Jumbo insect trap (Great Lakes IPM, Inc., Vestaburg, MI), were tested to determine if they are able to transmit SBMV and BPMV. The beetles were given a 24-h acquisition access (individually in plastic petri dishes) to detached trifoliolate leaves of Phaseolus vulgaris 'Black Valentine' systemically infected with either BPMV or SBMV. Beetles that had fed on the infected bean leaves (as evidenced by feeding damage to the leaves) were caged individually in clear plastic cups on healthy 'Black Valentine' bean seedlings for a 24-h transmission access period. The test plants were grown in the greenhouse for 2 weeks after which they were evaluated for virus infection. In the first trial, BPMV was not transmitted (0 of 11 bait plants with feeding damage), whereas SBMV was efficiently transmitted (6 of 6 bait plants with feeding damage) as determined by agar-gel double-diffusion serology of test plants using polyclonal antibodies to SBMV and BPMV. In a second trial, with a different set of beetles, the transmission rate for BPMV was 3 of 46 (6.5%) and 33 of 43 (77%) for SBMV, and the test plants were evaluated for virus infection using Protein A ELISA. The higher transmission efficiency of SBMV by Japanese beetles when compared with BPMV is characteristic of these two viruses when they are transmitted by other leaffeeding beetles (1). These results are in contrast to an earlier report that Japanese beetles were not able to transmit BPMV from infected soybeans to healthy soybeans in Illinois (3). To our knowledge, this is the first report that the Japanese beetle, an introduced insect with a wide host range in the United States, is a vector of plant viruses.

HYPERIMMUNISATION OF RABBITS WITH ULTRASOUND DISINTEGRATED BORDETELLA BRONCHISEPTICA AND CONTROL OF ANTIBODY SYNTESIS TROUGH DOUBLE DIFFUSION AGAR GEL

Two rabbits were hyperimmunisated with an ultrasound lysate of a pig origin B. bronchiseptica and one rabbit was hyperimmunisated with an ultras ound lysate of B. bronchiseptica dog strain origin. Each rabbit was inoculated three times. Blo od samples were collected before the hyperimmunisation begun and after each inoculation. Antibody producti on during immunization was controlled using the method of double diffusion in agar gel. After the first inocu lation only 2 of 3 rabbits responded with antibody production detectable with this method, but after the third in oculation all the rabbits responded with antibody p roduction at least against three antigenic compounds of the ultr asound disintegrated B.bronchiseptica . Serum samples obtained from each rabbit after the third antigen i noculation were tested against both antigen solutio ns. These tests proved that all rabbits produced antibodies a gainst the same three antigenic compounds of B. bronchiseptica.

동부(Vigna sinensis)의 국부병반을 이용한 Cucumber mosaic virus와 Broad bean wilt virus 2의 구별

Cucumber mosaic virus(CMV) and Broad bean wilt virus 2(BBWV2) were isolated from Gentiana scabra plants showing typical mosaic and yellowing symptoms. When the inoculum of mosaic symptom propagated in Nicotiana benthamiana was inoculated to primary leaves of Vigna sinensis, the local lesions of different types was developed. Type one produced a small necrotic spot(SNS) of pinpoint type, while the other one showed a large necrotic spot(LNS) of halo type. LNS on primary leaves of V. sinensis was also induced by inoculum from yellowing symptom on G scabra. Single lesion from SNS induced a typical mosaic symptom on N. Benthamiana. On the other hand, LNS produced a chlorotic ring symptom on inoculated leaves and mosaic plus necrotic ringspot on upper leaves of N. benthamiana. An isolate of CMV from SNS and BBWV2 from LNS were detected by using dsRNA analysis, RT-PCR and agar gel double-diffusion test. Thus, our results should provide a tool of a simple method for discrimination from mixed infected plants by CMV and BBWV2.

Identification of Tomato mosaic virus Infection in Lisianthus in Taiwan

In recent years, Lisianthus (Eustoma russellianum (Don.) Griseb) has become popular as potted plants and cut flowers in Taiwan. They are grown in the central and southern regions of the island. Since 1998, diseased plants with mosaic symptoms, followed by necrosis of leaf tissues, were observed in commercial greenhouses and field-grown lisianthus. Newly emergent leaves were curled and smaller compared with those on healthy plants. These symptoms greatly decreased the commercial value of the crop. Rigid rods similar to tobamoviruses that measured 300 × 18 nm were found consistently associated with symptomatic plants. In July 2002, a virus culture was isolated from diseased lisianthus from Chiayi County, Taiwan and established and maintained in systemic hosts Nicotiana tabacum L. and N. benthamiana Domin. Chlorotic and necrotic spots developed on lisianthus leaves 1 to 2 weeks after inoculation with the virus; symptoms eventually became systemic. Virions were purified from inoculated N. tabacum. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that the virus contained one 18-kDa (Mr) polypeptide. The virus reacted positively in agar gel double diffusion tests and enzyme-linked immunosorbent assays with antisera prepared to Tobacco mosaic virus (TMV) or Tomato mosaic virus (ToMV) (gifts of S. D. Yeh, National Chung Hsing University, Taichung, Taiwan). A viral coat protein (CP) gene approximately 0.5 kb was amplified by reverse transcription-polymerase chain reaction from total RNA prepared from infected N. benthamiana using 5'-GCGAGCCATGGATTCTTACTCAATTACT as a forward primer and 5'-ACTCTCGGATCCTTAAGATGCAGGTGCAGA as a reverse primer. Comparison of the 480-nt CP gene region with that of ToMV-OM (GenBank Accession No. X02144) (3) revealed 99.2% nucleotide identity and 99.4% amino acid identity. It shares, however, 74.4% nucleotide identity and 83.9% amino acid identity with CP genes of TMV-U1 (GenBank Accession No. AX040174) and TMV-vulgare (GenBank Accession No. J02415) (1). The virus induced local lesion responses similar to ToMV on inoculated N. tabacum cv. White Burley, N. sylvestris Speg. & Comes, and Datura stramonium L. Inoculation of TMV, however, resulted in a systemic infection in these plants. Results from sequence analysis and diagnosis based on host reaction to the virus inoculation indicated that the tobamovirus infecting lisianthus in Taiwan is an isolate of ToMV. The virus is economically important to lisianthus and tomato in Taiwan. To our knowledge, this is the first report of ToMV causing disease on lisianthus in Taiwan. The disease was previously observed on lisianthus in Italy (2).

Immunodiagnosis of ABPA.

For the diagnosis of allergic aspergillosis demonstration of specific immune responses to allergens has been accepted as a significant paradigm. Elevated levels of total IgE and Aspergillus fumigatus specific IgE and IgG antibodies are important criteria for diagnosis of ABPA. Although reference antigens or standardized methods are not available, there are a number of relevant recombinant antigens, which have been isolated in recent years. Several techniques have been employed in the demonstration of specific antibodies against antigens of Aspergillus fumigatus in the sera of patients. Of these methods, the widely followed ones are ELISA, radioimmunoassay, Western blot, and agar gel double diffusion. Recently, semi-automated methods have been developed using recombinant allergens to detect circulating antibody against Aspergillus. However, these methods have not been evaluated widely. Here we review the immunodiagnostic methods currently in use for allergic bronchopulmonary aspergillosis.

Identificação de predadores de Orphulella punctata (de Geer) (Orthoptera, Acrididae) através da serologia

Females of Orphulella punctata (de Geer, 1794) were captured in the field and kept in cages with food and boxes filled with moist soil for oviposition. The eggs, macerated with 0,85% saline solution and centrifugated, were used as the immunizing antigen for obtention of the specific antiserum (AS-O). Rabbits were immunized by the linphonodule injection method with two injections of this antigen, with an interval of 15 days between the 1st and 2nd inoculations. Serological tests were performed using double diffusion in agar gel with homologous and heterologous antigens. Specific serological reactions were obtained 14 days after the 1st inoculation. The antiserum of the 30th day gave 5, 2-4 and 1-4 precipitation lines respectively with eggs, female and male antigen. Such differences can be attributed to the high specificity of AS-O. Positive reaction was observed with arachnids and mirmeleontids captured in the field, indicating the predation on O. punctata.

Anti-topoisomerase I (Anti-Scl-70) antibodies in patients with systemic lupus erythematosus

To investigate the presence and clinical significance of anti-Scl-70 antibodies in patients with systemic lupus erythematosus (SLE). Levels of antibodies against Scl-70 were determined by a commercial clinical enzyme-linked immunosorbent assay (ELISA) during routine evaluation. Results were verified by an additional ELISA with a characterized bovine Scl-70, by ELISA with a recombinant human topoisomerase I, by Western blot, and by double diffusion in agar gel. Disease activity was estimated retrospectively by the Systemic Lupus Activity Measure (SLAM). Of 128 consecutive SLE patients, 25% were positive for anti-Scl-70 antibody; this antibody activity was cognate in nature. No SLE patient could be classified as also having systemic sclerosis. The levels of anti-Scl-70 were significantly correlated with the SLAM score for the entire cohort (r = 0.563, P < 0.001) and for 7 individual patients with multiple longitudinal measurements (r = 0.755-0.951, P < 0.001; n = 6) (r = 0.378, P < 0.05; n = 1). A significant correlation was also found between the levels of anti-Scl-70 and anti-double-stranded DNA antibodies (r = 0.558, P < 0.001). Patients with anti-Scl-70 had significantly higher risk of pulmonary hypertension (P < 0.01) and renal involvement (P < 0.001) than patients without this antibody. Anti-Scl-70 antibody is present in a significant subset of patients with SLE. For this subset, it offers a good correlate of disease activity and suggests increased risk for pulmonary hypertension and nephritis.

Serological properties of Abiotrophia and Granulicatella species (nutritionally variant streptococci).

Serological variations were examined among 12 type or reference strains and 91 oral isolates of vitamin B6-dependent Abiotrophia and Granulicatella spp. Rabbits were immunized with whole cells of 12 selected strains and 10 typing antisera were obtained, which were unreactive with the Lancefield group A to G antigen preparations. The reactivity of the antisera and autoclaved cell surface antigen extracts was tested by double diffusion in agar gel and a capillary precipitin test. These typing antisera categorized all Abiotrophia defectiva strains, all except one Granulicatella elegans strain, three-quarters of the Granulicatella adiacens, and half of the Granulicatella paraadiacens into 8 serotypes and 2 subserotypes. The Granulicatella balaenopterae type strain was unserotypable. All A. defectiva strains were serotype I, some of which were divided into subserotype I-1 and/or I-5. The G. adiacens strains generally belonged to serotype II or III, and the G. paraadiacens strains to serotype IV, V or VI. All G. adiacens or G. paraadiacens serotype II strains were also subserotype I-5. The G. elegans strains were serotype VII or VIII. These Abiotrophia and Granulicatella serotypes were undetectable among 33 strains of the other 11 species including the bacteriolytic enzyme-producing but vitamin B6-independent strains of Streptococcus, Enterococcus, Dolosigranulum and Aerococcus. The proposed serotyping system for Abiotrophia and Granulicatella spp. would be helpful in the identification and classification of these unique coccal isolates in ecological and epidemiological studies.

Comparative Analysis of the Effect of the VaccinesTALOVAC 109 SE and SG VACCINE NOBILISin the Formation of Immunity Against Salmonella Enteritidis

Two groups of 60-day-old chickens of Hisex white breed were respectively vaccinated with inactivated TALOVAC 109 SE and live SG VACCINE NOBILIS. Four months after vaccination, blood sera tests of both experimental groups were performed using the enzime immunoassay(ELISA) and reactions of double diffusion in agar gel with antigens specific for salmonellae. By ELISA there were defined relative values S/N of antibodies against Salmonella enteritidis as well as titre means in both groups. Experimental data show that in the first group vaccinated with TALOVAC 109 SE the mean of S/N values was 0,47 of relative units, whereas in the second group vaccinated with SG VACCINE NOBILIS S/N mean was lower and equaled 0,54. Applying double diffusion in agar gel method, a common precipitation link was found in both groups. It indicates, that the antigen against which antibodies developed in the organisms of chickens is common for S. enteritidis and S. gallinarum pathogens.

Uso de serologia na identificação de predadores de Orphulella punctata (De Geer) (Orthoptera: Acrididae)

Serological techniques were used to study Orphulella punctata (De Geer) predators. For preparation of antiserum, rabbits were immunized by the limphonodule injection method with antigens obtained from O. punctata male. Serological tests were performed using double diffusion in agar gel. Specific serological reactions were obtained 13 days after the 1st inoculation. After the 2nd inoculation 5 precipitation lines were observed. In the reactions with female antigens there were 2-4 lines, while egg antigens showed 2 lines. These variations in the number of lines were attributed to different physiological states and to ontogeny. Positive reactions were observed for spiders and chrysopids captured in the field.