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Adherence of Helicobacter pylori to gastric epithelial cells is thought to be important in the pathogenesis of infection and may be essential to maintain lifelong colonization. However, the factors responsible for adherence to gastric epithelial cells in vivo have not been characterized, and the significance of adherence to standard epithelial cell lines is unclear. Hemagglutination is also thought to be important in H. pylori adherence. However, no studies have clearly linked H. pylori hemagglutination or adherence to cultured epithelial cells to primary gastric epithelial cell adherence. Furthermore, it is not clear whether laboratory strains which have undergone multiple passages lose potential colonization factors. In this study, we examined the effect of serial laboratory passage on hemagglutination and correlated the hemagglutination characteristics of H. pylori strains to primary gastric cell adherence. Variable expression of hemagglutination was seen with serial laboratory passage of 15 strains. After 100 serial laboratory passages, all strains had lost hemagglutination activity. Hemagglutination was seen in association with adherence to primary gastric cells in vitro isolated from 2 patients. An association with ultrastructural intimate adherence was seen with HEp-2 cells, but not with gastric adenocarcinoma cells. Ultrastructural adherence was seen in corresponding antral biopsies of patients whose strains were hemagglutination positive, but hemagglutination was not associated with gastric inflammation. These data indicate that H. pylori hemagglutination is lost with serial passage and that hemagglutination may play a role in the attachment of H. pylori to gastric epithelial cells, but the role of adherence to chronic gastric inflammation is unclear.
Comparative evaluations of immune status for rubella virus are described for enzyme-linked immunosorbent assay, hemagglutination inhibition, and indirect hemagglutination. A 92.1% agreement between enzyme-linked immunosorbent assay and indirect hemagglutination assay was demonstrated for rubella immune status. Enzyme-linked immunosorbent assay and hemagglutination inhibition demonstrated a 92.6% agreement and were compared in an attempt to define the quantitative usefulness of comparisons of single sera for determining immune status. These data support the relative lack of correlation between single enzyme-linked immunosorbent assay and hemagglutination inhibition quantitative values. Enzyme immunoassay was, however, an acceptable alternative to hemagglutination inhibition for the determination of immune status to rubella virus.
The ability of Mycoplasma hyopneumoniae to agglutinate RBC was evaluated to develop an in vitro cytadsorption assay. Using swine RBC in a microtitration hemagglutination test, no agglutination or partial agglutination was detected. Comparison of RBC from various other species indicated that improved hemagglutination was obtained with RBC from turkeys. This hemagglutination was detected only when mycoplasma cells used in the assay had been frozen and thawed, heated at 50 C for 30 minutes, or treated with trypsin. Treatment of RBC with trypsin or neuraminidase enhanced hemagglutination. Possible surface lectin activity in M hyopneumoniae was evaluated by use of carbohydrates in a blocking assay; hemagglutination was not inhibited by any of 13 carbohydrates evaluated. Mycoplasma hyopneumoniae convalescent porcine serum and monoclonal antibodies against 2 M hyopneumoniae immunogens of molecular weights of 64,000 and 41,000 inhibited hemagglutination.
Standardized hemagglutination and hemagglutination-inhibition procedures are described and statistically evaluated for all animal viruses where applicable, except for rubella and the arbovirus group. The standardized tests employ a constant phosphate-buffered saline diluent and constant volumes of serum, antigen, and standardized erythrocyte suspension. The standardized hemagglutination test has a reproducibility of 84 to 96% with adenoviruses, rubeola, and the myxoviruses, and 78 to 93% with reoviruses; the standardized hemagglutination-inhibition test has a reproducibility of 95 to 100% with all viruses tested.
Various concentrations of pig, guinea pig, chicken, and turkey erythrocytes (RBCs) were cryopreserved and compared with fresh RBCs in hemagglutination and hemagglutination inhibition tests. Hemagglutination and hemagglutination inhibition titers obtained with cryopreserved or fresh RBCs from the same donor differed by not more than one twofold dilution. RBCs cryopreserved at a concentration of less than or equal to 12.5% gave the highest percent recovery upon thawing.
The experiments with the collection of strains and diagnostic sera available in the USSR confirmed the data contained in the literature on the good prospects of using the indirect hemagglutination test for the serotyping of bacteria belonging to the genus Providencia. Certain differences were observed in cross reactions, revealed by the indirect hemagglutination test and by agglutination: the interrelations of serogroups O14 and O25, O23 and O25, O23 and O29, O29 and O45, O37 and O49, determined by means of the indirect hemagglutination test, could not be detected in the same sera by the slide and tube agglutination tests. For the first time the usefulness of aqueous-saline extracts of agar cultures in the indirect hemagglutination inhibition test to specify more precisely the interrelations between individual bacterial serogroups of the genus Providencia has been shown.
The antigens were prepared from three B. thetaiotaomicron strains of different origin (NCTC 10582--reference strain, 312/85--isolated from a pancreas fistula drain of a patient, and 9/18--derived from normal human intestinal microflora). Lipopolysaccharides were extracted by the hot phenol-water method and purified by nuclease tretment. Degradation of lipopolysaccharides was achieved by mild acid hydrolysis obtaining polysaccharide (PS) and lipid (LA) fractions. Capsular polysaccharide (CPS) was extracted from the clinical strain 312/85 producing thick capsules. Antibacterial sera were prepared by immunization of rabbits with formolized suspensions of investigated strains. HA and IHA were performed with all antigens and immune sera. Examined cell-surface antigens (LPSs and CPS) were capable of coating formolized sheep erythrocytes. The titres obtained in the passive hemagglutination test in homologous reactions were 160-320. Polysaccharide fractions (PS) prepared by means of mild acid hydrolysis of LPSs were unable to coat formolized sheep red blood cells. The activity of sensitization of sheep erythrocytes was revealed by lipid fractions of LPSs. All preparations were active as inhibitors in the inhibition of passive hemagglutination. The strongest inhibitors in homologous systems were polysaccharide fractions of LPSs and capsular polysaccharide (the concentration of inhuibitor 8-15 micrograms/ml). The results of performed serological tests indicated the antigenic similarity of standard B. thetaiotaomicron strain NCTC 10582 and clinical strain 312/85. The strain 9/18 from normal human intestinal microflora showed distinct antigenicity. High-molecular cell-surface B. thetaiotaomicron antigens containing carbohydrates (LPS and CPS) can be applied in the passive hemagglutination test and in the inhibition of passive hemagglutination.
Like enteroviruses, hepatitis A virus (HAV) hemagglutinated various species of erythrocytes under similar conditions. HAV-specific antibodies in both acute- and convalescent-phase sera were found to inhibit hemagglutination. The HAV hemagglutination inhibition test can be used for diagnosis, epidemiological surveillance, and vaccine assessment.
Erythrocytes collected from 1-day-old chicks were stabilized by fixation with formaldehyde and by freeze-drying after treatment with carbon monoxide. Suspensions of freeze-dried erythrocytes in distilled water or physiological saline had a homogeneous bright reddish-purple color. Freeze-dried erythrocytes were compared with fresh erythrocytes for hemagglutination and hemagglutination-inhibition tests for various viruses including rubella, Japanese encephalitis, influenza, mumps, Newcastle disease, and Sendai viruses. After storage at 4 degrees C for 1 year or more, freeze-dried erythrocytes maintained their original appearance and sensitivity to hemagglutination antigens.
Conditions for psittacine beak and feather disease (PBFD) virus hemagglutination and hemagglutination-inhibition (HI) test reactions are defined. The PBFD virus was found to hemagglutinate cockatoo and some guinea pig erythrocytes. The HI test was used to assay serum antibody titer in birds with active PBFD virus infections and in others that had been exposed to diseased birds. On the basis of HI antibody titers in psittacine birds that had been exposed to PBFD virus, but remained clinically normal, we suggest that some birds exposed to the virus are able to mount an effective immune response. Birds with active PBFD virus infections had lower antibody values than did birds that had been exposed to the virus, but remained clinically normal. On the basis of these findings, the ability to develop a suitable HI antibody response may be crucial in determining the disease status of susceptible birds exposed to the PBFD virus. If HI antibodies are found to have neutralizing activity, then the fact that a high HI titer was induced in birds inoculated with purified PBFD virus might suggest that an immunization program would be effective in preventing PBFD virus infections.
Newly developed serological methods for the detection of pili in the passive hemagglutination (PHA) test with the use of immunoglobulin erythrocytic diagnosticum and in the enzyme immunoassay (EIA) with the use of specific immunoglobulins labeled with horse radish peroxidase have been found to exceed the method of detecting pili, based on the determination of their hemagglutinating activity, in sensitivity and specificity. Besides, the PHA test and EIA have proved to be capable of detecting low molecular fragments of pili, obtained by sonication and having lost their hemagglutinating activity.
The HEPES-saline-albumin-gelatin (HSAG) diluent found optimal for agglutination of fowl erythrocytes by rubella virus antigen is also optimal for agglutination of trypsin-treated human group O cells. Albumins from different commercial sources, however, can have varying inhibitory effects on rubella hemagglutination titers. This can have a significant effect on the hemagglutination-inhibition test since antibody titers measured by this procedure are related to the amount of antigen used.
Chuzan virus agglutinated erythrocytes of several species of animals including bovine. The hemagglutinating (HA) activity against bovine erythrocytes was dependent on NaCl molarity and was expressed best at 0.6 M, but it was independent of pH and temperature. Three strains of Chuzan virus isolated from 2 cows and a pool of culicoides midges had indistinguishable HA antigenicity. All cattle infected with the virus developed high titers of hemagglutination inhibiting (HI) antibody which changed in parallel with neutralizing (NT) antibody titers. Correlation between HI and NT antibodies was very high and the antibodies persisted for one year or more. Therefore it was concluded that the HI test is applicable for survey of Chuzan virus infection among cattle in place of the NT test.