Agglutination Tests ; methods ; Brucellosis ; blood ; diagnosis ; Coombs Test ; methods ; Humans ; Sensitivity and Specificity

Due to the serovar diversity in Haemophilus (H.) parasuis, it is difficult to develop a universal serological method for detection of this pathogen. Here, we report a universal plate-agglutination test for detecting H. parasuis. Diagnostic antisera were prepared by mixing antisera of serovars 4, 5, 12, 13 and 14 in the optimized ratio. The results of the plate-agglutination test showed that the diagnostic antisera could agglutinate with all 15 reference strains of H. parasuis and 74/75 clinical isolates. Further, the specificity of the method was validated with 22 bacterial strains from 12 related species.
Agglutination Tests/*methods ; Animals ; Cross Reactions ; Haemophilus parasuis/isolation & purification/*physiology ; Immune Sera/*metabolism ; Reproducibility of Results ; Sensitivity and Specificity

OBJECTIVETo evaluation the specificity and sensitivity of 5 kinds of serological detection methods about brucellosis.

METHODSTo investigate in the 4 autonomous banner (Cha You Hou Qi, Right-Wing Central Banner of Kerqin Region, Linxi County and Siziwangqi Banner) of Inner Mongolia autonomous region from January to December, 2013. Accepting criteria: professionals of breeding cattle and sheep, and slaughter,accompanied by Bloom's disease suspected symptoms such as fever, fatigue,arthralgia, ranging in age from 25 to 55 years old. To collect suspected patients venous blood 3-5 ml in the morning, a total of 236 samples were collected. To detect the Brucella antibody by using plate agglutination test (PAT), tiger red plate agglutination test (RBPT), standard test tube agglutination test (SAT), enzyme-linked immunosorbent assay (ELISA) and immune colloidal gold method (GICA), SAT was taken as a golden standard, analyzed the sensitivity and specificity of RBPT and SAT, ELISA and GICA.

RESULTSSAT method of positive patients: 136 cases (57.6%). PAT method positive patients: 150 cases (63.6%). RBPT positive patients: 159 cases (67.4%), and 143 patients with ELISA method: positive (60.6%), 147 patients with positive GICA method (62.3%). The detection rate of Brucella antibody positive was different by different testing methods.There was no significant difference (χ(2)=0.52,P=0.264). To take the SAT method as the gold standard, PAT, RBPT, ELISA and GICA method of the sensitivity were 97.7% (133/136), 98.5% (134/136), 94.8% (129/136) and 94.1% (128/136), respectively. The specificity was lower,the rate were 70.0% (70/100), 75.0% (75/100), 86.0% (86/100) and 81.0% (81/100), respectively. The total coincidence rate were 86.0% (203/236), 88.5% (209/236), 91.1% (215/236) and 88.5% (209/236), respectively.

CONCLUSIONThe specificity and sensitivity of ELISA and GICA method is higher in the diagnosis of disease. The two methods are rapid, GICA method can be used on-site testing, large sample test is suitable for using ELISA.

Adult ; Agglutination Tests ; methods ; Animals ; Antibodies, Bacterial ; blood ; Brucella ; Brucellosis ; diagnosis ; Cattle ; China ; Enzyme-Linked Immunosorbent Assay ; Humans ; Middle Aged ; Sensitivity and Specificity ; Sheep

BACKGROUND: We compared the results of automated and quantitative methods for the diagnosis of syphilis, Mediace Rapid Plasma Reagin (RPR) and Mediace Treponema pallidum Latex Agglutination (TPLA) (Sekisui Chemical Co., Ltd, Japan) with those of conventional methods. METHODS: Sera from 3,896 persons who had health checkups between December 2005 and November 2006 were included in the evaluation of positive rates and biological false positives (BFP) for Mediace RPR and TPLA. In addition, 134 patients' sera positive for automated Mediace RPR or TPLA were tested for VDRL and TPHA. Discrepancies between TPLA and TPHA results were confirmed by the RecomBlot Treponemal IgG/IgM (Mikrogen GmbH, Germany). Automated Mediace RPR and TPLA were performed using the Hitachi 7600 chemistry autoanalyzer (Hitachi, Japan). Samples with positive Mediace RPR and negative TPLA results were defined as BFP. RESULTS: Positive rate of automated Mediace RPR was 0.23% (9/3,896). BFP of the Mediace RPR was 0.18%. Positive rate of automated TPLA was 1.62% (37/2,284). Among the 134 patients' sera, 33 (24.6%) showed a discrepancy between conventional VDRL and automated Mediace RPR results: Among 31 Mediace RPR(+)/VDRL(-) sera, 13 were positive and 18 were negative for TPLA. The remaining 2 sera of discrepancy with Mediace RPR(-)/VDRL(+) were all positive for TPLA. There were seven sera that showed a discrepancy between automated TPLA and TPHA results: Two sera with Mediace RPR(+)/TPLA(-)/TPHA(+) showed negative recomBlot Treponemal IgG/IgM results, and among five sera with TPLA(+)/TPHA(-), three demonstrated IgG or IgM by recomBlot Treponemal IgG/IgM. CONCLUSIONS: The results of comparison data demonstrated that automated TPLA results had a high concordance with recomBlot Treponemal IgG/IgM results. Moreover, there are additional advantages of automated methods such as quantitative detection, low infection risk, and no influence by human handling.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Agglutination ; False Positive Reactions ; Female ; Humans ; Immunoglobulin G/analysis ; Immunoglobulin M/analysis ; Latex Fixation Tests ; Male ; Middle Aged ; Reagent Kits, Diagnostic ; Reagins/*blood ; Syphilis/*diagnosis ; Syphilis Serodiagnosis/*methods ; Treponema pallidum/*immunology/isolation & purification

One hundred seven isolates of Staphylococcus aureus from bovine mastitis were investigated for colony morphology in serum-soft agar (SSA), autoagglutination in salt, and capsular serotype. Capsular polysaccharide (CP) was purified and quantified from the extracts of clinical isolates. Overall, 89 isolates (83.2%) were diffuse in the SSA, without any difference in the proportion of diffuse colony between type 5 and type 8 strains. Some strains exhibited compact colonies in the SSA and expressed CP as determined by an enzyme-linked immunosorbent assay, indicating that compact morphology does not exclude encapsulation. The majority of the strains (11/12) showed autoagglutination in the salt aggregation test. The serotype 336 accounted for 46.7% of the isolates followed by serotype 5 (12.1%) and serotype 8 (12.1%). Particularly, twenty-six (24.3%) isolates reacted with two serotypes; 7 for type 8/336 and 19 for type 5/336. Five isolates (4.7%) were nontypeable with monoclonal antibodies specific for CP serotype 5, 8, or 336. The CP concentration in culture supernatants varied with the serotypes, and the total amount of CP produced by cells grown in a liquid medium was much less than that produced by cells grown on a solid medium. The Western blotting indicated that the CP bands of S. aureus serotype 5 and 8 were ranged in the molecular mass of 58-84 kilodalton (kDa), with additional bands in the region of approximately >or= 48 or Agglutination Tests ; Animals ; Bacterial Vaccines/administration & dosage ; Cattle ; Cattle Diseases/immunology/microbiology ; Female ; Mastitis, Bovine/*microbiology ; Polysaccharides, Bacterial/*classification/isolation & purification ; Serotyping/methods/veterinary ; Staphylococcal Infections/immunology/*veterinary ; Staphylococcus aureus/*classification/*immunology/isolation & purification

OBJECTIVETo establish ELISAs based on rLipL32/1-LipL21-OmpL1/2 fusion antigen of Leptospira interrogans for detecting specific IgG and IgM in serum of patients with leptospirosis.

METHODSMicroscope agglutination test(MAT) was performed to detect serum specimens from leptospirosis patients and to determine titers of rabbbit antiserum agaist rLipL32/1-LipL21-OmpL1/2 to reference standard strains of L. interrogans. By using rLipL32/1-LipL21-OmpL1/2, rLipL32/1, rLipL21 and rOmpL1/2 as the coated antigens, ELISAs for detecting specific serum IgM and IgG were established. The established ELISAs were applied to MAT-positive serum specimens from 107 patients with leptospirosis.

RESULTThe results of MAT confirmed that 66% (71/107) of the patients were infected with L.interrogans serogroup Icterohaemorrhagiae, and the rLipL32/1-LipL21-OmpL1/2 antiserum were able to agglutinate all 15 reference standard L.interrogans strains with 1 : 20approximate, equals1 : 160 titers. The positive rates of ELISAs using rLipL32/1-LipL21-OmpL1/2, rLipL32/1, rLipL21 or rOmpL1/2 as the antigen were 89.7%, 75.7%, 85.1% and 79.4% for detecting IgM, respectively, while 99.1%, 99.1%, 94.4% and 86.0% for detecting IgG, respectively. The positive detection rate of rLipL32/1-LipL21-OmpL1/2-IgM-ELISA was higher than those of the other three IgM detection ELISAs (P<0.05). The positive detection rate of rLipL32/1-LipL21-OmpL1/2-IgG-ELISA was higher than that of rOmpL1/2-IgG-ELISA (P<0.05), while there was no significant differnce with that of rLipL21-IgG-ELISA and rLipL32/1-IgG-ELISA (P>0.05).

CONCLUSIONThe ELISAs using rLipL32/1-LipL21-OmpL1/2 as the antigen can be applied as a sensitive,specific and universal serological method for diagnosis of leptospirosis.rLipL32/1-LipL21-OmpL1/2-IgM-ELISA shows a definite value for early diagnosis of leptospirosis compared with the other ELISAs used in this study.

Agglutination Tests ; Animals ; Antibodies, Bacterial ; blood ; Antigens, Bacterial ; genetics ; metabolism ; Bacterial Outer Membrane Proteins ; genetics ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; genetics ; metabolism ; Humans ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Leptospira interrogans ; genetics ; immunology ; Leptospirosis ; diagnosis ; immunology ; Lipoproteins ; genetics ; metabolism ; Rabbits ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Sensitivity and Specificity ; Serologic Tests ; methods

Escherichia coli (E. coli) strains were collected from young diarrheic calves in farms and field. Strains that expressed the K99 (F5) antigen were identified by agglutination tests using reference antibodies to K99 antigen and electron microscopy. The K99 antigen from a selected field strain (SAR-14) was heat-extracted and fractionated on a Sepharose CL-4B column. Further purification was carried out by sodium deoxycholate treatment and/or ion-exchange chromatography. Monoclonal antibodies to purified K99 antigen were produced by the hybridoma technique, and a specific clone, NEK99-5.6.12, was selected for propagation in tissue culture. The antibodies, thus obtained, were affinity-purified, characterized and coated onto Giemsastained Cowan-I strain of Staphylococcus aureus (S. aureus). The antibody-coated S. aureus were used in a coagglutination test to detect K99+ E. coli isolated from feces of diarrheic calves. The specificity of the test was validated against reference monoclonal antibodies used in co-agglutination tests, as well as in ELISA. Specificity of the monoclonal antibodies was also tested against various Gram negative bacteria. The developed antibodies specifically detected purified K99 antigen in immunoblots, as well as K99+ E. coli in ELISA and co-agglutination tests. The co-agglutination test was specific and convenient for large-scale screening of K99+ E. coli isolates.
Agglutination Tests/methods/*veterinary ; Animals ; *Animals, Newborn ; Antibodies, Monoclonal/*immunology ; Antigens, Surface/immunology/isolation & purification ; Bacterial Toxins/immunology/isolation & purification ; Cattle ; Cattle Diseases/*immunology/*microbiology ; Chromatography, Gel/veterinary ; Chromatography, Ion Exchange/veterinary ; Chromatography, Liquid/veterinary ; Diarrhea/immunology/*veterinary ; Electrophoresis, Polyacrylamide Gel/veterinary ; Enzyme-Linked Immunosorbent Assay/veterinary ; Escherichia coli/*immunology ; Escherichia coli Infections/immunology/*veterinary ; Immunoblotting/veterinary ; Staphylococcus aureus