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1.
Two new monoclonal antibodies (MAbs), D6D8D5 and B3E6F9, both directed against Haemophilus paragallinarum serovar C hemagglutinating (HA) antigen, were produced, and characteristics of the MAbs were compared with those of the previously described MAb F2E6 in dot-blot and hemagglutination-inhibition (HI) tests using two representative H. paragallinarum strains each of serovars A, B, and C strains and 55 Japanese serovar C field isolates. MAb D6D8D5 and MAb F2E6 reacted with all serovar C strains and field isolates in the dot-blot test. However, MAb D6D8D5 showed various degrees of inhibition of the HA activity of field isolates. In the enzyme-linked immunosorbent assay-competition test, MAb D6D8D5 did not compete with MAb F2E6. MAb B3E6F9 reacted with strain S1, serovar C but not with strain Modesto, serovar C in both dot-blot and HI tests. Three out of 55 field isolates did not react with MAb B3E6F9. Neither MAb reacted with the serovar A and B strains.  相似文献   

2.
Two monoclonal antibodies (MAbs), lMAb-1 and lMAb-5, against Actinobacillus pleuropneumoniae serotype 1 were obtained. In enzyme-linked immunosorbent assay-inhibition tests with whole cell antigens obtained from serotype 1 to 12 strains of A. pleuropneumoniae, lMAb-1 reacted to only a serotype 1, strain 4074. The epitope recognized by lMAb-1 was a carbohydrate sensitive to periodate oxidation and resided on capsular polysaccharide (CP) of A. pleuropneumoniae serotype 1. On the other hand, lMAb-5 reacted with serotype 1, 9 and 11 strains at the same degree and its epitope was found to be located on O-polysaccharide of serotype 1, 9 or 11 lipopolysaccharide (LPS). These results showed that CP was one of the serotype-specific antigens of A. pleuropneumoniae, and that O-polysaccharide of LPS obtained from serotype 1, 9 or 11 strain was the cross-reacting antigen among these strains.  相似文献   

3.
A panel of four monoclonal antibodies (MAbs) was evaluated, using a hemagglutination-inhibition test, for its ability to subtype 76 isolates of Haemophilus paragallinarum. The results of the MAb reactions were compared with the results of both the Page and Kume serotyping schemes (the serovars of the Page scheme correspond to the serogroups of the Kume scheme). One MAb (E5C12D10) was raised against a Page serovar A strain and the remaining MAbs (F2E6, D6D8D5, and B3E6F9) against a Page serovar C strain. Six different reaction patterns were found among the 76 isolates of H. paragallinarum. There was total correlation between the MAb reaction pattern and the Page scheme, and thus the Kume scheme, to the serogroup level. All 19 Page serovar A (= Kume serogroup A) strains reacted only with MAb E5C12D10, whereas all five Page serovar B (= Kume serogroup B) strains failed to react with any of the MAbs. All 52 remaining strains were Page serovar C (= Kume serogroup C), and all failed to react with MAb E5C12D10 but showed varying reaction patterns with the three other MAbs. Although the MAbs recognized four subdivisions within Kume serogroup C, these subdivisions differed from the four Kume C serovars. This panel of MAbs can be used to assign isolates of H. paragallinarum to either Page serovars or Kume serogroups. Although the subdivisions recognized by the MAbs within the Page serovar C strains do not correspond to the Kume serovars, they may be useful in epidemiological applications.  相似文献   

4.
S Y Kang  L J Saif 《Avian diseases》1991,35(3):563-571
Fifteen monoclonal antibodies (MAbs) against an avian group A rotavirus were cloned and characterized. Eight of the 15 MAbs had neutralizing activity (N-MAbs). Five of the N-MAbs (1G1, 5B8, 4E2, 3G1, 2E3) were VP4-specific by radioimmunoprecipitation assay (RIPA), and two N-MAbs (2D11, 6E8) were possibly VP7-specific (faint bands by RIPA). One N-MAb (4H12) of undefined protein specificity cross-reacted with serotype 3 simian rotaviruses. The other seven N-MAbs did not cross-react with any of the eight distinct serotypes of human and mammalian rotaviruses tested. Of the seven non-neutralizing MAbs, three were VP6-specific (3H10, 4B12, 5F6), two were VP8-specific (6C9, 1D1), one was VP4-specific (4E9), and one was of undefined protein specificity (1B11). Four non-neutralizing MAbs recognized only avian group A rotavirus in cell-culture immunofluorescence tests (6C9, 1D1, 4E9 and 5F6), whereas two MAbs (3H10 and 4B12) cross-reacted with all human and animal rotaviruses tested. The MAb 1B11 did not recognize any human rotavirus serotypes but cross-reacted with all nonhuman animal rotavirus serotypes. The MAbs produced in this study should be useful for the detection and further characterization of avian group A rotaviruses.  相似文献   

5.
抗副鸡嗜血杆菌血清A和C型株所制备的两个血清型单克隆抗体（MAbs），分别对副鸡嗜血杆菌血清型A、B、C中的各型参考株作HI和dot－blotting试验。一种MAb（E5C12D10）为抗血清型A代表株221，另一种MAb（F2E6）为抗血清型C代表株S1。在两种试验中，不同血清型的MAbs可与对应的血清型中的副鸡嗜血杆菌株血凝（HA）抗原反应，而与血清型B代表株91、147均无反应。故这些MAbs可用于dot－blotting或HI试验进行副鸡嗜血杆菌定型。  相似文献   

6.
Two serovar-specific monoclonal antibodies (MAbs) to Haemophilus paragallinarum serovars A/1 and C/2 strains, respectively, were developed and characterized by hemagglutination-inhibition (HI) and dot-blotting tests using representative H. paragallinarum serovars A/1, B, and C/2 strains. In both the HI and dot-blotting tests, one MAb (E5C12D10), raised against strain 221, serovar A/1, reacted only with serovar A/1 strains, while the other MAb (F2E6), raised against strain S1 of serovar C/2, reacted with only serovar C/2 strains examined. In both tests, the two MAbs did not react with two serovar B strains. These results indicated that the two MAbs recognize serovar-specific hemagglutinating (HA) antigens of H. paragallinarum serovars A/1 and C/2 strains, respectively, and that a dot-blotting test using these MAbs is a practical alternative to the HI test for serotyping H. paragallinarum. Strains 0222 and Spross of serovar B, which did not react with these two MAbs, were found to possess serovar-specific HA antigen in cross-HI tests.  相似文献   

7.
The antigenic differences between strains of serotype 2 of both biotypes I and II of Actinobacillus pleuropneumoniae were studied by using several serological techniques. Monoclonal antibodies (MAbs) against A. pleuropneumoniae biotype I serotype 2 were produced by fusion of spleen cells of BALb/c mice immunized with whole-cell (WC) suspension with SP2/O-Ag14 murine myeloma cells. Desirable MAbs were selected by enzyme-linked immunosorbent assay (ELISA) using WC as antigen. MAbs MK-7 and MK-10 identified multiple bands of lipopolysaccharide in Western-blot. Treatment of WC with proteinase K and sodium periodate indicated that both MAb binding sites were carbohydrates in nature. In both ELISA and Western-blot, MAbs MK-7 and MK-10 recognized only biotype I serotype 2 isolates. Neither MAb MK-7 nor MK-10 reacted with reference strains of remaining serotypes of A. pleuropneumoniae and other Gram-negative bacteria tested. The results obtained with various serological tests showed that strains of serotype 2 biotype I shared antigenic determinants with strain N-282 of serotype 2 biotype II, but not with strain N-273 of serotype 1 biotype II. It is suggested that data obtained from this study may be helpful in the development of specific serotyping and serodiagnostic reagents of A. pleuropneumoniae strains.  相似文献   

8.
Five nonneutralizing monoclonal antibodies (MAb) generated to the virulent Miller strain of transmissible gastroenteritis virus (TGEV) and specific for the S protein were characterized. Competition assays between purified and biotinylated MAb indicated that MAb 75B10 and 8G11 mapped near a new subsite, designated V and 2 MAb, 44C11 and 45A8, mapped to a previously designated subsite D. A fifth MAb mapped between subsites V and E. These MAb were tested with 3 previously characterized MAb to subsites A, E, and F in fixed-cell ELISA and cell culture immunofluorescent assays against 5 reference and 9 field strains of TGEV and 2 US strains (ISU-1 and ISU-3) of porcine respiratory coronavirus (PRCV). Subsites A, E, and F were conserved on all TGEV and PRCV strains examined. The 2 MAb to subsite V, 8G11 and 75B10, reacted only with the Miller TGEV strains (M5C, M6, and M60), except that 75B10 also recognized field strain U328. The MAb 11H8 did not react with 4 field strains or the Purdue strains of TGEV. The 2 MAb to subsite D reacted with all TGEV strains examined, but not with 2 US PRCV strains, 2 European PRCV strains, 1 feline infectious peritonitis virus strain, and 1 canine coronavirus strain. Because of this specificity for TGEV, but not PRCV, these latter 2 subsite D MAb may be useful for the development of competition ELISA to differentiate serologically between TGEV and PRCV infections in swine, similar to the currently used European subsite D MAb.  相似文献   

9.
The kinetics of the antigen production of Chlamydia psittaci strains Izawa-1 and Pigeon-1041 (P-1041) was examined every 6 hr after infection up to 48 hr, by the indirect immunofluorescent antibody technique using monoclonal antibodies (MAbs). All three genus-specific antigenic determinants on lipopolysaccharide (LPS) appeared during the whole growth cycle. Antigenic determinants on proteins were, on the other hand, detected at various time periods from the early to the late stages of infection. However, a cross-reactive antigenic determinant on protein recognized by a MAb 3E9 was also detected during the whole growth cycle, similar to that on LPS. The time of appearance of common antigenic determinants on proteins of Izawa-1 and P-1041 was examined using cross-reactive MAbs, and it varied depending on heterologous and homologous MAbs. From the relationship between the detection of antigenic determinants and the morphological changes of chlamydial particles revealed by electron microscopy during the growth cycle, the antigenic determinants on proteins of Chlamydia psittaci were divided into two groups; one was specific to the elementary body and the other was coexisting in both the elementary body and the reticulate body.  相似文献   

10.
Three antigens were prepared from Pasteurella multocida strain P-1059, and their immunogenicity and antigenic relationships were investigated. The 3 antigens were a soluble antigen purified from a 2.5% NaCl extract (2.5S), a similar antigen purified from an extract in 0.3% formalin solution containing 0.85% NaCl (FS), and lipopolysaccharide (LPS). The antigens were treated with various chemicals and enzymes to study their antigenic and immunogenic determinants. Antigenic analyses with ELISA inhibition tests indicated that 2.5S and FS were similar LPS-protein complex antigens. The 2.5S and FS antigens induced protective immunity in turkeys with high antibody titers against LPS antigen. Although LPS was a component of 2.5S and FS, LPS itself was poorly immunogenic in turkeys. The antigenicity of protein compounds in 2.5S was deteriorated by protease treatment, which, however, did not significantly diminish the protective immunogenicity. Treatment of 2.5S with sodium periodate, altering its carbohydrate moieties, decreased its immunogenicity. The immunogenicity of 2.5S also was abolished by phenol-water treatment, owing to dissociation of the LPS-protein complex. These findings suggest that a certain form of LPS-protein complex is essential for the induction of immunity against the P multocida infection in turkeys.  相似文献   

11.
Two monoclonal antibodies (MAbs) were evaluated for their ability to serotype 108 isolates of Haemophilus paragallinarum. One MAb (E5C12D10) was raised against a Page serovar A strain and the other (F2E6) against a Page serovar C strain. In both dot blot and hemagglutination-inhibition tests, MAb E5C12D10 recognized the type strains of Page serovar A and Kume serovars A-1, A-2, A-3, and A-4. MAb F2E6 recognized the type strains of Page serovar C and Kume serovars C-1, C-2, and C-3. Neither antibody recognized the type strains of Page serovar B or Kume serovars B-1 and C-4. When evaluated with 97 field isolates in a dot blot test, the MAbs serotyped 81 isolates, which was better than agglutinin typing by the Page scheme (69 isolates serotyped). The field isolates that did not react with the MAbs were either Page serovar B/Kume serovar B-1 (three isolates), Page serovar C/Kume serovar C-4 (12 isolates), or nontypable by either the Page or Kume scheme (one isolate).  相似文献   

12.
Four hybridoma cell lines producing monoclonal antibodies (MAbs) against Actinobacillus (Haemophilus) pleuropneumoniae were established by fusion of mouse myeloma and spleen cells obtained from mice immunized with a serotype 2, strain SH-15. Enzyme-linked immunosorbent assay-inhibition tests with antigens obtained from 12 serotype strains of A. pleuropneumoniae and 9 other gram-negative bacteria showed that all the MAbs bound to only serotype 2 strains of A. pleuropneumoniae. The epitopes recognized by the MAbs were located on a carbohydrate moiety of lipopolysaccharide (LPS) of the organism, which was sensitive to periodate oxidation. In immunoblotting analyses of LPS obtained from A. pleuropneumoniae serotype 2, all the four MAbs reacted specifically with the characteristic ladder bands of LPS detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These results suggest that O-antigen side chains of the LPS are one of the antigenic determinants responsible for the serotype-specificity of A. pleuropneumoniae.  相似文献   

13.
Edwardsiella tarda is an important cause for hemorrhagic septicemia in fish and gastro and extra-intestinal infections in humans. Monoclonal antibodies (MAbs) were produced against outer membrane proteins (OMPs) of E. tarda ET-7, isolated from diseased snakehead (Ophiocephalus punctatus). Two stable hybridoma clones, designated as 3F10 and 2C3 MAbs were found to be potentially specific for E. tarda by indirect enzyme linked immunosorbent assay (ELISA). These MAbs recognized major immunogenic OMP band at 44kDa in Western blotting. Both MAbs belonged to the IgG1 isotype and recognized different epitopes of OMP as seen by competitive ELISA. These MAbs strongly reacted with all 17 isolates of E. tarda used in our study by indirect ELISA and Western blotting. Interestingly, no reaction was observed with the reference strain of E. tarda (MTCC 2400). The sensitivity of 3F10 MAb to detect whole cells of E. tarda was up to a level of 1x10(4)CFU/ml in indirect ELISA. No cross-reactivity of MAbs were seen with Escherichia coli, Salmonella arizonae, Pseudomonas fluorescens, Aeromonas hydrophila, Vibrio cholerae, Flavobacterium ferrugineum and Mycobacterium tuberculosis. These MAbs could be used for specific detection of E. tarda infection in fish by immunoassays.  相似文献   

14.
A simple colony immunoblotting method using monoclonal antibodies (MAbs) was developed to detect Y. enterocolitica serotype O:3 in pig feces. One of the MAbs studied was able to detect single colonies of the organism in the presence of calculated 3.1 x 10(8) heterologous organisms in pig feces. The MAb was found to be specific for the lipopolysaccharide (LPS) O-antigens of Y. enterocolitica serotype O:3. No significant cross-reactivity was found against a variety of closely related serotypes and Gram-negative organisms. The MAb could also be used in a slide agglutination test and an indirect fluorescence antibody assay for rapid identification of Y. enterocolitica serotype O:3.  相似文献   

15.
A Qin  L F Lee  A Fadly  H Hunt  Z Cui 《Avian diseases》2001,45(4):938-945
In an attempt to develop a specific diagnostic test for avian leukosis virus (ALV) subgroup J (ALV-J) strain Hc1, four monoclonal antibodies (MAbs), JE9, G2, 145, and J47, were generated that are specific for ALV-J envelope glycoprotein, gp85. Polymerase chain reaction (PCR) was used to amplify genomic pro-viral DNA of Avian Disease and Oncology Laboratory (ADOL)-Hc1 and ADOL-4817 envelope genes. Both open reading frames encoding glycoproteins gp85 and gp37 were cloned into baculoviruses. Abundant expression of gp85 and gp37 was detected in the recombinant viruses with specific antibody to Hc1 strain of the ALV-J. The expressed proteins were used for immunization of mice to produce hybridoma cell lines secreting MAbs specific to ALV-J envelope protein. A panel of MAbs was generated by fusing NS1 myeloma cells and spleen cells from mice immunized with the recombinant baculoviruses. With the use of an immunofluorescence assay, three MAbs (JE9, G2, 145) reacted with ALV-J but not with subgroups A, B, C, D, or E of ALV. MAb J47 reacted with all exogenous subgroups of ALV including A, B, C, D, and J but not with endogenous subgroup E viruses. Western blot analysis was performed with all four MAbs against recombinant baculovirus and Hc1-infected chicken embryo fibroblast (CEF) lysates. A major band with a molecular weight about 90 kD corresponding to the size of ALV-J envelope was consistently obtained. With these MAbs, we detected the Hc1 antigen in CEFs infected with several ALV-J viruses isolated in the United States and also in tissue sections from chickens infected with Hc1 strain of ALV-J. These MAbs will be useful reagents for the diagnosis of ALV-J infection because they recognize a common antigenic epitope in six isolates tested thus far.  相似文献   

16.
Western blots of proteins of 14 Mycoplasma iowae strains and isolates resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were probed with three monoclonal antibodies (MAbs), MI6, MI7, and MI8. MAb MI6 reacted with one or more antigens with apparent molecular weights of 60,000, 70,000, and 94,000. In three strains (N-PHN-D13, R-D2497, and K 1805), antigens located on a single peptide band were recognized, while in others additional epitopes at different molecular-weight positions were revealed. A similar pattern was observed with MAb MI7, although it reacted with fewer antigens than did MAb MI6 and failed to recognize antigens in strains N-PHN-D13 and R-D2497. MAb MI8 reacted with an antigen at an apparent molecular-weight position of 28,000 in four of the 14 strains and isolates. The diverse reaction patterns observed with the MAbs in the 14 M. iowae strains and isolates confirms the occurrence of antigenic variation within this species. Antigenic variation in M. iowae may be pivotal in determining host-parasite interactions, pathogenesis, and the outcome of disease.  相似文献   

17.
Neutralizing epitopes present on field isolates of bluetongue virus (BTV) serotypes 10, 11, 13 and 17 were evaluated with a panel of polyclonal and neutralizing monoclonal antibodies (MAbs). A total of 91 field isolates were evaluated, including 15 isolates of BTV-10, 29 isolates of BTV-11, 26 isolates of BTV-13, and 21 isolates of BTV-17. The viruses were isolated from cattle, goats, sheep, elk and deer in Idaho, Louisiana, Nebraska and, predominantly, California, in the years 1979, 1980 and 1981. The isolates were analyzed and compared using a panel of neutralizing MAbs which included five MAbs raised against BTV-2, seven against BTV-10, five against BTV-13, and six against BTV-17. Neutralization patterns obtained with the MAb panel and individual field isolates were compared to those obtained with prototype viruses of each serotype. All field isolates were neutralized by at least some of the MAbs raised against the prototype virus of the same serotype. All field isolates of BTV-10 were neutralized by the seven MAbs raised to BTV-10, whereas the field isolates of BTV-11, BTV-13 and BTV-17 were not consistently neutralized by all of the MAbs raised against the prototype virus of the same serotype. Variation in neutralizing epitopes recognized by the MAb panel was most pronounced amongst the field isolates of BTV-17. A one-way cross neutralization was evident between BTV-10 and BTV-17 as all field isolates of BTV-17 were neutralized by four of the MAbs raised against BTV-10. In contrast, no BTV-10 isolates were neutralized by the MAbs raised against BTV-17. Differences in the MAb neutralization patterns of field isolates of BTV-11, BTV-13 and BTV-17 suggest that the immunogenic domain responsible for their neutralization is plastic, such that individual epitopes within the domain may vary in their significance to the neutralization of different viruses, even of the same serotype. The apparent conservation of neutralizing epitopes on field isolates of BTV-10 suggests that the field isolates may be derived from the modified-live vaccine strain of BTV-10.  相似文献   

18.
Vaccines for the prevention of fowl pox in chickens and turkeys have been available for more than five decades. However, in recent years outbreaks have occurred in several previously vaccinated chicken flocks. Presumably, fowl poxviruses (FPVs) antigenically different from the attenuated vaccine strains are responsible for such occurrences. In support of this concept, we previously detected minor antigenic changes in field isolates based on comparative immunoblotting with polyclonal anti-FPV serum. Realizing the need for antibodies specific against the dominant antigens of FPV, monoclonal antibodies (MAbs) were produced by immunizing mice with either a field strain of FPV or a pigeon poxvirus, currently used for vaccination. Three hybridoma clones producing MAbs reacting with a specific FPV protein were selected from a total of 83 clones. In immunoblots, two of the MAbs, P1D9 and P2H10, recognized an antigen with an apparent molecular weight varying from 39 to 46 kD, depending on the FPV strain. The third MAb, P2D4, reacted with an approximately 80-kD protein, regardless of which FPV isolate was tested. Immunofluorescent staining with P1D9 and P2D4 revealed that these MAbs react with intracytoplasmic antigens in FPV-infected cells.  相似文献   

19.
We prepared 31 monoclonal antibodies (MAbs) against either FIPV strain 79-1146 or FECV strain 79-1683, and tested them for reactivity with various coronaviruses by indirect fluorescent antibody assay (IFA). Sixteen MAbs which reacted with all of the 11 strains of feline coronaviruses, also reacted with canine coronavirus (CCV) and transmissible gastroenteritis virus (TGEV). In many of them, the polypeptide specificity was the recognition of transmembrane (E1) protein of the virus. We succeeded in obtaining MAbs which did not react with eight strains of FIPV Type I viruses (showing cell-associated growth) but reacted with FIPV Type II (79-1146, KU-1) and/or FECV Type II (79-1683) (showing non-cell associated growth). These MAbs also reacted with CCV or TGEV. These MAbs recognized peplomer (E2) glycoprotein, and many antigenic differences were found in this E2 protein. These results suggest that FIPV Type II and FECV Type II viruses are antigenically closer to TGEV or CCV than to FIPV Type I viruses. Furthermore, the MAb prepared in this study has enabled discrimination between FIPV strain 79-1146 and FECV strain 79-1683, which was thought to be impossible by the previous serological method.  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   

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