首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献，搜索用时 421 毫秒
1.
The polypeptides of serologically related viruses of hemorrhagic enteritis (HE) in turkeys, marble spleen disease (MSD) in pheasants, and splenomegaly in chickens (SMC) were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by protein immunoblotting with polyclonal antibodies to HE virus (HEV). The viral polypeptides II, III, IV, V, VI, and VII were detected on SDS-PAGE with the size range from 18 to 97 kDa in HEV. Viral polypeptides II, III, V, VI, and VII were detected in MSD virus and virus of SMC. Protein immunoblotting of viral proteins with anti-HEV serum revealed antigenic differences between the 3 viruses of avian adenovirus type-II examined. The differences were that the polypeptides II, III, IV, V, VI, and VII were identified in HEV and the polypeptides II, V, VI, and VII were identified in MSD virus and virus of SMC. The bands of penton base (polypeptide III) and fiber (polypeptide IV) were seen in HEV only by protein immunoblotting.  相似文献   

2.
The present study was undertaken to antigenically characterize the buffalopox virus (BPV). Six monoclonal antibodies (MAbs) against the BP4 strain of BPV have been produced and characterized. All six MAbs appeared to be specific to BPV, as none of them showed cross-reactivity with other poxviruses in antigen capture ELISA. Only two MAbs (20AB8 and 20CD11) bound significantly with different BPV isolates in antigen capture ELISA, whereas the remaining four MAbs bound weakly with the BPV. In Western blot analysis with purified BPV-BP4, the rabbit hyperimmune serum against purified BPV-BP4 reacted with 15 immunodominant polypeptides (100 kDa to 25 kDa), whereas two MAbs (21CB6, 21DB11) reacted with 42 kDa and 45 kDa polypeptides, respectively. However, three MAbs (20AB8, 20CD11, 21CB5) reacted with three degraded polypeptides (100 kDa, 40 kDa and 87 kDa) of BPV-BP4. In radioimmunoprecipitation assay (RIPA) with the rabbit hyperimmune serum to BPV-BP4, three virus specific polypeptides (69 kDa, 34 kDa, 32 kDa) were recognized in BPV-BP4, whereas two polypeptides (69 kDa, 34 kDa) were recognized in other BPV isolates (BPV-Bly, BPV-Vij96, BPV-Vij97). In virus neutralization test, none of the six MAbs tested showed any significant neutralizing ability to infection with different BPV isolates. However, the hyperimmune serum showed weak neutralizing ability to BPV infection.  相似文献   

3.
Outer membrane proteins (OMP) of P. multocida (serotype B:2) field isolates (n = 6) and a vaccine strain (P-52) were extracted by a sarkosyl method and characterized using SDS-PAGE and immunoblotting. About 20 polypeptide bands were observed in the profile of the vaccine strain with MW ranging from 16 to 90 kDa and, based on band thickness and intensity of staining, three polypeptides of MW 31, 33 and 37 kDa were considered to be the major OMPs. The profiles of the field isolates showed minor differences when compared with that of the vaccine strain. The OMP of 33 kDa was only expressed by the vaccine strain. Four field isolates expressed an OMP of 39 kDa, which did not appear in the profiles of the remaining two field isolates and the P-52 strain. Similarly, an OMP of 25 kDa was exclusively seen in the profile of a single isolate. By immunoblotting studies, using anti-P. multocida (P-52) whole-cell hyperimmune serum raised in rabbits as well as buffalo immune sera, it became evident that the polypeptide of 37 kDa was the most antigenic OMP in the profiles of all the isolates, including the P-52 strain. Other polypeptides were either weakly antigenic or visible in the profile of only a few of the isolates. The study thus identified the major OMP of P. multocida (B:2) and suggested that this highly antigenic 37 kDa OMP has potential for further protective and immunodiagnostic studies.  相似文献   

4.
This study was carried out to identify immunoreactive polypeptides in Babesia equi merozoite antigen. Three fractions of killed B. equi merozoite antigen viz.; whole merozoite (WM), cell membrane (CM) and high speed supernatant (HSS) antigens were prepared from the parasite infected erythrocytes. These antigenic preparations along with ghost antigen from non-infected erythrocytes were fractionated on 12% sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotted with sera showing high antibody titres. On SDS-PAGE, 16 polypeptides with molecular weight (Mr) in the range of 112-17kDa were obtained from the WM and CM antigens. But only six polypeptides were detected (96.5-28kDa) in the HSS antigen. On immunoblotting with high titred serum collected from donkeys following two immunizations with a killed B. equi merozoite immunogen, 11 polypeptides were observed in the WM and CM antigens (Mr 112-18kDa). Of these, four polypeptides (Mr 112, 45, 33 and 18kDa) were identified as most immunoreactive. Besides these, a 28kDa was observed as strong immunoreactive protein in WM and CM antigens. The HSS antigen showed only six polypeptides and one peptide (28kDa) was identified as immunoreactive. When high titred serum collected from immunized donkeys following challenge with B. equi infected blood and was used for immunoblotting, the protein profile of WM and CM antigens remained the same. However, three additional polypeptides (Mr 81, 54.5 and 39kDa) were detected in HSS antigen.  相似文献   

5.
Ewe placental and lamb intestinal isolates of Chlamydia psittaci recovered from flocks affected with ovine enzootic abortion were examined by inclusion morphology, indirect immunofluorescence (IIF) and immunoblot analysis. Chlamydiae recovered from the faeces of sheep from two flocks free of clinical disease were also examined. In cell culture ovine abortion (OA) and intestinal isolates were distinguishable by inclusion development and morphology. Similarly, in two-way IIF tests with one week mouse antisera isolates fell into two distinct groups: abortion or intestinal. Immunoblotting with convalescent sheep abortion antiserum identified 30 out of at least 40 silver staining polypeptides as antigenic both in OA and intestinal isolates. The serum produced a similar reaction pattern to the resolved proteins of each OA isolate, indicating a higher degree of antigenic conservation among these isolates. Considerable cross reactivity between the OA and intestinal isolates was identified, but the serum also showed apparent molecular weight differences between antigens of the two types in the 87-116 kDa, 38-44 kDa and 26-28 kDa regions. Furthermore, the immunoblotting analysis revealed heterogeneity among the intestinal isolates, particularly in antigens between 87-116 kDa and 38-44 kDa.  相似文献   

6.
A trivalent antigen for the detection of type I avian adenovirus precipitin   总被引：1，自引：0，他引：1  
B S Cowen 《Avian diseases》1987,31(2):351-354
A double immunodiffusion antigen prepared from cell-culture-propagated CELO virus was not capable of detecting precipitin directed against all of the type I avian adenovirus (fowl adenoviruses) isolates tested. However, an antigen pool containing CELO-4, B-3, and IBH-2 (Tipton) fowl adenovirus isolates detected precipitin directed against representative isolates of 10 type I serotypes. Additionally, this antigen pool markedly improved detection of adenovirus field exposure.  相似文献   

7.
Monoclonal antibodies were produced against orf virus-specified cell surface proteins in an attempt to develop reagents capable of differentiating between members of the Parapoxviridae. Two immunization protocols were used to induce an anti-orf response in BALB/c mice, one of which resulted in virus replication in the recipient. The monoclonal antibodies produced were tested for crossreactivity with bovine papular stomatitis virus (BPS) and milker's node virus (MNV) by indirect immunofluorescence assay (IFA) and immunoblotting. The results indicate that significant antigenic overlap exists between isolates of orf, MNV and BPS, even at the level of specificity provided by monoclonal antibodies. One monoclonal antibody reacted strongly in IFA with orf virus isolates, very weakly with MNV, and not at all with BPS. On immunoblots this same antibody recognized a 40-43 kDa protein in orf virus-infected cells, and also a 45-48 kDa protein in cells infected with MNV or BPS virus. The data suggest that it may be possible to define parapoxvirus strains on the basis of small variations in specific virus-directed cell surface proteins.  相似文献   

8.
The antigenic polypeptides in Salmonella Enteritidis (SE) were chronologically recognized by the chicken immune system, using Western immunoblotting. Broiler chicks challenged at three days of age with SE strain carrying the most prevalent plasmid profile of 14.1 and about (approximately) 50 kb were bled at 17, 24, 31, 38 and 45 days of age. Pooled sera of blood collected at each age was reacted by Western immunoblotting with banded polypeptides of three predominant SE strains that acquired the following respective plasmid profiles: 14.1 kb; 14.1 and approximately 50.0 kb; and 1.8, 14.1 and approximately 50.0 kb. The immunoblots of each pooled sera collected at a specific age against the three SE strains were similar. More specifically, the early immune response at 17 days of age had antibodies recognizing only one polypeptide in the three SE strains namely, the 35.8 kDa. At 24 or 31 days of age, the acquired immunity to infection had antibodies recognizing five similar polypeptides in the three SE strains namely, the 14.4 (fimbriae protein), 21.5 (fimbriae protein), 30.5, 35.8, and 66.2 kDa. At 38 and 45 days of age, the antibodies recognized additional polypeptides namely, the 41.5 and 55.6 kDa, respectively. The recognition of the 41.5 and 55.6 kDa polypeptides at 45 days of age was associated with higher invasiveness of SE to spleens and livers (15.6%) and in higher cecal colonization (59.4%) in comparison to absence of recognition to the two polypeptides at 31 days of age associated with low infectivity to spleens (0%), livers (3.1%), and ceca (9.4%).  相似文献   

9.
Both avian adenovirus-associated virus (A-AV) and CELO virus were isolated from the embryonating eggs of 25-week-old black sex-linked hens during a naturally occurring infection. In the first 7 days of egg collection, A-AV was isolated from 10 of 43 (23.2%) embryonating eggs, and CELO virus was isolated from 8 of 43 (18.6%) embryonating eggs. Both viruses were isolated from six eggs. In the next 16 days of egg collection, A-AV and CELO virus were coisolated from 1 of 127 (0.8%) eggs; all other samples were negative for both viruses. All six hens transmitting A-AV to eggs and 5 of 6 hens transmitting CELO virus showed seroconversions (fourfold increase in antibody concentrations). Viruses were not isolated from eggs after the hens showed a fourfold increase in antibody concentrations.  相似文献   

10.
The protein profiles of Pasteurella multocida serotype 1 isolates and the response of chickens to serotype 1 antigens were investigated using SDS-PAGE. Patterns obtained with Coomassie blue staining of soluble protein extracts were similar. The major difference between isolates was the position of one of the major proteins in the 34-38 kDa region. When chickens were experimentally infected with a clinical isolate of P. multocida serotype 1 various proteins were recognised by immunoblotting, including one with a relative molecular weight of 34 kDa; however, no reactions were observed in the region where LPS is known to migrate. When these infection sera were used in an EIA with purified LPS obtained from Heddleston serotype 1 type strain (X-73) they reacted strongly. Serum used for serotyping isolates in the gel diffusion precipitin test recognised many antigens in common with sera from infected birds, but some antigens were specific to typing sera.  相似文献   

11.
The cytoplasmic protein antigens (CPAg) of Brucella canis were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and analysis of 35S-labeled polypeptides. Approximate molecular weights of the immunoreactive polypeptides were determined by migration patterns of the immunoprecipitated polypeptides after SDS-PAGE or Western immunoblotting of sera collected at various times after experimental infection of dogs. Polypeptides were specifically precipitated by sera of infected dogs, but not from the sera of normal or false-positive (seropositive, non-infected) animals. During the initial month after infection, proteins with molecular weight masses (MW) of approximately 18, 22, 31, 42 and 54 kDa were commonly recognized. A 20-kDa polypeptide was first recognized at 8-10 weeks after infection, but it was detected inconsistently after 6 months. Additional polypeptides detected from 2 to 12 months post-infection had MW of 22, 66-68 and, less regularly, 42, 60, 82, 100 and greater than 200 kDa. The polypeptides most consistently recognized in sera from B. canis-infected dogs had MW of 18, 22 and 68 kDa.  相似文献   

12.
The aim of this study was to use the enzyme-linked immunosorbent assay (ELISA) and the Western immunoblotting as possible tools to differentiate infections in turkeys by different paramyxoviruses. Pooled hyperimmune sera of turkeys infected with either paramyxovirus-3 (PMV-3), paramyxovirus-6 (PMV-6), or Newcastle disease virus (NDV) were assayed for antibodies specific to the three viruses by the ELISA and Western immunoblotting. ELISA results showed cross reactions of turkey antibodies between PMV-3 and PMV-6 antigens, while turkey antibodies to NDV did not cross-react with any of the other paramyxoviruses. The immunoblots of sera from birds infected with PMV-3 (Minnesota turkeys and Iowa chickens) reacted to low molecular weight polypeptides of PMV-3 of 29, 32, and 34 kDa, and to a high molecular weight band of 200 kDa. The same Minnesota turkey sera had a cross reaction to the 200 kDa polypeptide of PMV-6, while the Iowa chicken sera did not. Both sera had no apparent reaction to NDV proteins. Western immunoblotting showed that the turkey PMV-3 sera had a specific reaction to a 220 kDa polypeptide present in PMV-3, but not in PMV-6, while the turkey PMV-6 sera had a specific reaction to a 130 kDa polypeptide present in PMV-6, but not in PMV-3. Immunoblots of pooled sera from turkeys infected with PMV-6 (Minnesota source) reacted to the 200 kDa protein present in both PMV-3 and PMV-6; however, no reaction occurred between this sera and NDV proteins.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

13.
Y Okuda  M Ono  S Yazawa  Y Imai  I Shibata  S Sato 《Avian diseases》2001,45(4):819-827
The pathogenicity of a serotype 1 fowl adenovirus (FAV-99ZH), isolated from broiler chickens exhibiting gizzard erosion, was investigated in commercial broiler chickens. Five-, 3-, and 1-wk-old commercial broiler chickens were inoculated with FAV-99ZH by both oral and ocular routes. In the 5-wk-old chickens (trial 1), none of which had the maternal antibody to FAV-99ZH, severe gizzard erosions were observed on days 5, 7, and 10 postinoculation (PI). Among the 3-wk-old chickens (trial 2), which were separated into a control group and three treatment groups according to their maternal antibody titer levels, some chickens showed clinical signs such as depression and anorexia. Compared with the control group, all the treatment groups showed decreased weight gain. One treatment group, moreover, showed significantly decreased (P < 0.05) weight gain on day 10 PI. Severe gizzard lesions, such as erosion or ulcers, were observed from day 4 PI in all treatment groups regardless of their maternal antibody levels. The 1-wk-old chickens (trial 3) were separated into a control group and two treatment groups according to their titer levels of the inoculated virus. In spite of high maternal antibody levels, severe gizzard lesions were observed in both treatment groups, which also showed decreased weight gain. One treatment group, inoculated with the higher dose, showed significantly decreased (P < 0.05) weight gain on days 10 and 14 PI compared with the control group. Fowl adenovirus was recovered mainly from gizzard and rectal (including feces) samples from inoculated chickens but was not recovered from liver samples in any of the trials or in any of the control chickens. Although the reproduced disease was similar to that described in a previous report of experimental infection of specific-pathogen-free (SPF) white leghorn chickens with fowl adenovirus, the pathogenicity of FAV-99ZH in commercial broiler chickens was more severe than that in the SPF white leghorn chickens. The results of the present study indicate that FAV-99ZH isolated from gizzard erosion had pathogenicity and produced severe lesions in the gizzards of broiler chickens and that FAV-99ZH could infect and produce illness in broiler chickens with maternal antibodies against this virus.  相似文献   

14.
为了克隆禽腺联病毒(Avian adeno-associated virus,AAAV)全基因组用于构建基因转移载体研究,以鸡胚致死孤儿病毒(CELO)作为辅助病毒与AAAV共接种SPF鸡胚进行AAAV的增殖,将AAAV约4.7kb双链基因组DNA与pCR2.1载体连接,构建了含AAAV全基因组的重组质粒pAAAV并进行了测序。序列分析表明,AAAV YZ-1株的基因组为4684bp,两端具有141bp的末端倒置重复序列和Rep蛋白结合位点特征序列,与GenBank中收录的AAAV DA-1株和VR-865株的核苷酸序列同源性分别为95.0%和92.2%。将pAAAV质粒转染CELO病毒感染的鸡胚肝细胞系,获得了感染性AAAV病毒粒子,结果证明克隆的AAAV基因组中存在与病毒复制和包装相关的正确关键序列,可用于重组AAAV载体的构建。  相似文献   

15.
Zoospores of Dermatophilus congolensis were analysed by SDS-PAGE and western blotting. The electrophoretic profiles of zoospores from 13 isolates of D. congolensis were similar but not identical when stained with Coomassie blue or silver. Immunodominant polypeptides with apparent molecular masses of 76 and 31 kDa were identified in western blots of 13 of 13 and 12 of 13 isolates respectively of D. congolensis reacted with hyperimmune, ovine, antizoospore sera. Identical immunodominant polypeptides were observed in western blots reacted with sera obtained from naturally infected sheep. Initial characterisation of the 76 and 31 kDa polypeptides indicated that they were probably surface exposed because (i) antibodies eluted from the surface of live zoospores after adsorption of hyperimmune antizoospore serum, reacted principally against the 76 and 31 kDa subunit polypeptides in western blots, (ii) adsorption of hyperimmune antizoospore serum with live zoospores resulted in significant diminution of reactivity against both the 76 and 31 kDa polypeptides in western blots, (iii) indirect fluorescent immunostaining of zoospores with antiserum prepared against gel-purified 76 kDa polypeptide, resulted in intense staining of the zoospore outer coat. Immuno-gold electron microscopy of negatively stained zoospores with antiserum prepared against gel-purified 31 kDa polypeptide identified this antigen as a flagella subunit.  相似文献   

16.
Four Theileria parva isolates from Muguga area of Kiambu district, Kenya, were used to establish schizont-infected cell lines. Their protein antigens were then separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS page). The isolates were subsequently subjected to protein analysis and characterisation by the western immunoblotting technique. Probing for the polymorphic immunodominant molecule (PIM) was done using monoclonal antibody no. 4. SDS page detected up to 20 protein antigens of molecular mass 35,000-180,000 Da. The western blot analysis revealed a greater heterogeneity in the molecular mass (M(r)) of PIM than previously thought. The M(r) of PIM varied between 80 and 90 kDa. The isolates further revealed different densities of surface epitopes with variable reaction to the monoclonal antibody. The implications of these findings to the epidemiology of east coast fever and immunisation programmes are discussed.  相似文献   

17.
The pathogenicity of a serotype-1 fowl adenovirus (FAV-99ZH), which causes adenoviral gizzard erosion by oral inoculation in chickens, was investigated in specific pathogen-free white leghorn chickens. In trial 1, 14 chickens were inoculated intravenously with the virus at 21 days of age and euthanatized for necropsy within 1-14 days of inoculation. Gizzard erosion was grossly observed from day 7 postinoculation (PI), and histologically, FAV-99ZH antigen-positive, basophilic intranuclear inclusion bodies were seen in the gizzard lesions from day 7 to 11 PI. Necrotizing pancreatitis, and cholecystitis and cholangitis associated with the inclusions were observed from day 3 to 14 PI (pancreatitis) and from day 5 to 9 PI (cholecystitis and cholangitis), respectively. The inclusions were also observed in the epithelial cells of the cecal tonsils from day 3 to 5 PI. The virus was recovered from samples of the lesions. It was revealed that FAV-99ZH causes not only gizzard erosion but also pancreatitis, cholecystitis, and cholangitis by intravenous inoculation in chickens. In trial 2, 10 chickens were inoculated orally with the virus twice, at 13 and 36 days of age, and euthanatized for necropsy within 4-17 days after reinfection. Macroscopically, focal gizzard lesions were observed; however, neither necrosis nor inclusions were observed by microscopy. Moreover, FAV was not recovered from the gizzard or rectum of any of the chickens at necropsy. This suggests that the gizzard lesions occurred as a result of the primary infection, and that the chickens were able to resist reinfection.  相似文献   

18.
Humoral responses were examined in rabbits immunized with either 28-40 kDa (Fraction 1) or a 19-24 kDa (Fraction 2) antigenic fraction from soluble antigens (Sol L3 Ag) from infective larvae (L3) of Haemonchus contortus. These fractions were eluted from electrophoretically separated Sol L3 Ag. Immunoblots revealed antibodies to Fraction 1 (fr. 1) or Fraction 2 (fr. 2) polypeptides as well as to several other molecular weight polypeptides of the Sol L3 Ag. The latter antibodies were shown by absorption studies not to be Sol L3 Ag cross-reactive anti-bacterial rabbit antibodies. When Sol L3 Ag was affinity-purified using monoclonal antibody to phosphorylcholine (PC) and the resulting fractions were further analysed by immunoblotting using rabbit anti fr. 1 or anti fr. 2 antiserum, the PC antigen was found to be shared between fr. 1 and other polypeptides of Sol L3 Ag. Using the rabbit antibody fractions eluted from nitrocellulose membranes containing fr. 1 or 2 polypeptides, it was found that these fractions contained antibody that bound mainly to fr. 1 and only to fr. 2 polypeptides of Sol L3 Ag. It is concluded that, from the present immune rabbit sera, antibodies specific for either fr. 1 or fr. 2 may be isolated and then used to purify small amounts of the corresponding antigens.  相似文献   

19.
Monoclonal antibodies (MAbs) were generated against the recombinant plasminogen activator (Pla) protein of Yersinia pestis. These MAbs detected Pla in all the 18 isolates of Y. pestis obtained from the sputum of pneumonic plague patients and from the liver and spleen of rodents from plague-affected areas of India during 1994-1995 as well as in seven of the eight isolates obtained from rodents in the surveillance regions of Hosur and Palmner in India during 1998 by simple dot-ELISA. In immunoblotting, the MAbs reacted with the Pla antigen only in Y. pestis isolates at 37 and 35kDa region. These monoclonal antibodies, being strictly specific, can be used for detecting Y. pestis isolates that are Fraction 1 antigen-negative. Also, the radiolabelled pla fragment hybridized specifically to the representative DNA samples of Y. pestis isolates.  相似文献   

20.
The use of 2 monoclonal antibodies (MAbs), P1D9 and P2D4, which recognize different fowlpox virus (FPV) antigens, for the identification and characterization of FPV strains was evaluated. Initially, the MAbs were used in conjunction with a dot blot assay that enabled FPV to be differentiated from the avian herpesvirus, infectious laryngotracheitis virus. Confirmation of the specificity of these MAbs was provided by the demonstration that only FPV antigens were recognized by a combination of both antibodies when used for immunoblotting proteins contained in various avipoxviruses. Later, an antigenic characterization of 11 FPV field isolates, 6 FPV vaccine strains, and 3 pigeonpox virus vaccines was performed by Western blotting with the individual MAbs. Whereas MAb P2D4 consistently recognized a protein with an apparent molecular weight of 60 kD, there was variability in the size of the antigen that was immunoreactive with the other MAb. For example, MAb P1D9 recognized an antigen of apparent molecular weight of 46 kD in all vaccine strains except 2 of FPV origin. In these exceptions, either only a 39-kD or both a 42- and 46-kD protein were immunoreactive. As for the field isolates, a 39-kD antigen was recognized in 8 of them, whereas a 42-kD antigen was detected in the remaining 3. Therefore, the more extensive immunoblotting technique may facilitate FPV strain differentiation, whereas routine diagnosis of fowlpox could be accomplished by using the MAb-based dot blot assay.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号