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A fraction from Swiss albino mouse brain homogenate obtained by DEAE cellulose chromatography showed a non-specific hemagglutination-inhibitory property for some Togaviruses. Evidence are presented indicating that the inhibitory property is related to a protein-rich fraction with sialic acid as an active group.
Heparin inhibited hemagglutination (HA) by pseudorabies virus (PRV), but not HA by Akabane virus, bovine adenovirus type 7, Fukuoka virus, Getah virus, Japanese encephalitis virus, and parainfluenza virus type 3 belonging to the families Bunyaviridae, Adenoviridae, Rhabdoviridae, Togaviridae, Flavivi-idae, and Paramyxoviridae, respectively. The minimal inhibitory concentration of heparin required to inhibit 8 HA U of PRV ranged from 0.005 to 0.01 U/ml. Mouse erythrocytes failed to combine with the HA inhibitory factor of heparin. On the other hand, mouse erythrocytes treated with heparinase had greatly reduced agglutinability by PRV. Virus-heparin complex formation could be observed by sedimenting heparin with the virus particles.
We studied long-term rubella hemagglutination inhibition (HI) antibody titers in 381 children born between 1965 and 1966 with congenital rubella syndrome and in 344 of their mothers, who had had postnatally acquired rubella infection. Serum specimens were obtained 12 times during a 17-year period. The initial geometric mean titer of the rubella HI antibody was higher in the children than in their mothers, but the titer declined more rapidly in the children. After 17 years it was 2(5.4) in the children and 2(6.9) in the mothers. In 13 (3.4%) of the children, the rubella HI antibody titer reverted to negativity after three to 17 years. None of the mothers showed a reversion.
Hemagglutination inhibition (HI), single radial hemolysis (SRH) and enzyme-linked immuno sorbent assay (ELISA), performed with commercial antigen and reagents are described and were compared in the three distinct situations that require rubella antibody detection. Determination of immunity status was carried out on 156 sera. A degree of correlation greater than 0.9 was found when comparing the three methods. Analysis of a further 74 sera, from 31 primary infections and three congenital syndromes, was performed to compare the occurrence of the various classes of antibodies in the three tests: HI test and IgM-ELISA become positive the day after the rash, whereas SRH test is not positive before the sixth day. From our limited study bearing on a total of 230 sera, each test has a precise assignment. For the determination of immunity status, SRH is simpler, faster, and inexpensive; absence or evidence of past infection can be unequivocally obtained especially in cases of low (1:10, 1:20) residual immunity. In the serodiagnosis of a rubella rash, SRH alone, due to the delayed rise in antibody titers, will demonstrate a complete seroconversion with a first serum collected up to the fifth day of the eruption. In case of absence of an early serum, of primary infection in a pregnant woman, of a newborn with suspicion of congenital syndrome, the measurement of rubella specific IgM is best obtained with ELISA, a procedure less time-consuming than HI following centrifugal, chromatographic, or electrophoretic separation, and "light" (8 S) RF with SRH test is discussed. Interference of IgM Rheumatoid Factor (RF) with IgM ELISA and IgG RF with SRH test is discussed.
Four methods to detect antibodies reactive with soluble nuclear antigens were compared-the fluorescent antibody technique, hemagglutination, complement fixation, and immunodiffusion. Centain advantages and disadvantages of each method are described. The first three methods have greater sensitivity than immunodiffusion, but immunodiffusion discriminates most reliably between different antibodies.
C3b receptor (CR1) on erythrocytes from 23 patients with systemic lupus erythematosus (SLE) and 124 normal controls was determined by immune adherence hemagglutination (IAHA) and radioimmunoassay. The binding of radiolabeled monoclonal anti-CR1 to erythrocytes and their lysate was distributed continuously in a wide range. The majority of SLE patients showed low binding by both assays. CR1 sites on erythrocytes were determined also by Scatchard plot analysis and standardized by the number of similarly determined lectin-binding sites that served as a measure of erythrocyte surface. The numbers of standardized CR1 sites were classified as high, intermediate, and low. Thirty-six percent of control subjects had high numbers of CR1 sites, 53% had intermediate numbers, and 11% had low numbers. Of SLE patients, the numbers of CR1 sites were high in 0%, medium in 52%, and low in 48%. Negative IAHA was found in 10 controls (8%), all of whom had low numbers of standardized CR1 sites. Among 13 SLE patients with negative IAHA, 11 had low numbers of CR1 sites and the remaining 2 had low intermediate numbers. IAHA, therefore, was particularly efficient in detecting the low numbers of CR1 sites in SLE, which would impair the disposal of circulating immune complexes and accelerate the development of tissue injuries.
The binding activity of polymerized human serum albumin was determined in 202 HBsAg carriers. The presence of polymerized human serum albumin receptor sites was tested by hemagglutination and differentiated from antihuman albumin antibodies by immunofluorescence, isolation of IgG and IgM fractions and testing of HBsAg anti-HBs immune complexes. A granular pattern with anti-HBs was specific for polymerized human serum albumin receptor sites as demonstrated with purified HBsAg. In addition, a linear pattern with fluoresceinated antihuman immunoglobulins might suggest the presence of antihuman albumin antibodies (which was generally due to an IgG antibody). However, a granular pattern with fluoresceinated antihuman immunoglobulins may indicate the presence of HBsAg anti-HBs immune complexes. A weak linear pattern was also observed simultaneously in these cases, probably due to IgM antihuman albumin antibodies or an antipolymerized human serum albumin receptor site antibody. Of 202 HBsAg-positive patients, 71 showed polymerized human serum albumin receptor sites activity. The highest percentage of polymerized human serum albumin receptor sites was found among patients showing HBeAg and hepatitis B virus DNA polymerase positivity (96%), followed by HBeAg positivity and hepatitis B virus DNA polymerase negativity (48%), and anti-HBe positivity and hepatitis B virus DNA polymerase negativity (17%). In addition, a significant correlation between polymerized human serum albumin titers and hepatitis B virus DNA polymerase was found (r = 0.573, p less than 0.01). However, at similar HBeAg titer, patients who were positive for hepatitis B virus DNA polymerase had a higher polymerized human serum albumin receptor sites titer than those who were negative for hepatitis B virus DNA polymerase.(ABSTRACT TRUNCATED AT 250 WORDS)
Fecal occult blood testing by immunochemical hemagglutination has been shown to be superior to the Hemoccult test, both in sensitivity and in specificity. The test has been widely used as a tool for population screening in Japan, but there has been no study to evaluate the efficacy of screening using this test. A case-control study to evaluate the screening was conducted in study areas where no previous and no other concomitant colorectal cancer screening had been performed. Case series in the study were 193 cases who died of colorectal cancer. Three controls were selected randomly from the list of individuals who were alive at the time of diagnosis of the corresponding case and had been living in the same area as the case, matched by gender and by age. Odds ratios (OR) of dying of colorectal cancer for those screened within 1, 2 and 3 years of case diagnosis vs. those not screened were 0.40 [95% confidence interval (CI) 0.17-0.92], 0.41 (95% CI 0.20-0.82), and 0.48 (95% CI 0.25-0.92), respectively. OR increased towards 1.0 as the duration during which screening histories were compared was extended, and showed similar tendencies when analyzed by number of years since the most recent screening history. These results suggest that colorectal cancer screening by the immunochemical fecal occult blood test would reduce mortality from colorectal cancer.
A 21% incidence of false-positive hemagglutination inhibition tests for pregnancy associated with tubo-ovarian abscess is described. Five illustrated cases and the causes for such false-positive reactions are discussed. Substances generated by pelvic inflammatory masses may interfere with immunologic assays for pregnancy, resulting in misdiagnosis or inappropriate management.
A new indirect hemagglutination (M-Toxo-IHA) test was standardized for the serodiagnosis of acute toxoplasmosis. The reagent for this test consisted of stabilized human red cells coated with a Toxoplasma gondii heat-stable alkaline-solubilized extract which reacted predominantly with IgM antibodies found in serum samples from patients with a recent infection. Almost no reactivity was verified with serum samples from chronic or ancient toxoplasmosis. The reagent preparation was simple and obviated prior fractionation of parasite components reactive with IgM antibodies. Rheumatoid factor or high levels of IgG antibodies did not interfere in the M-Toxo-IHA test. A total of 358 serum samples was assessed for the evaluation of this test, comprising sera from patients with recent and ancient toxoplasmosis, from non-related diseases, as well as from clinically healthy individuals with no previous T. gondii infections. Serodiagnostic performance of the M-Toxo-IHA test in terms of sensitivity, specificity, efficiency, and positive and negative predictive values was verified, and the values obtained were 1.000, 0.985, 0.989, 0.952, and 1.000, respectively, in relation to IgM-immunofluorescence or IgM-capture ELISA. Six different batches of reagent, successively produced for the current study, provided reproducible results and were stable for at least 22 months at 4 degrees C.
An immune adherence hemagglutination assay (IAHA) for the detection of antibody to the Norwalk agent of acute epidemic nonbacterial gastroenteritis was developed using as antigen virus purified from stool from an experimentally infected volunteer. The assay was sensitive and specific and was efficient for detecting Norwalk antibody seroresponses. The prevalence of Norwalk antibody in various groups in the United States was studied. Antibody to the Norwalk agent was acquired gradually, beginning slowly in childhood and accelerating in the adult period so that by the fifth decade 50% possessed antibody. This pattern of antibody acquisition contrasted sharply with that for the human rotavirus of infantile gastroenteritis. Rotavirus antibody was acquired during early childhood by almost all individuals in the pediatric groups studied. Antibody to the Norwalk agent was also found in rural Bangladesh; in a small prevalence survey of 39 children and adults 21% possessed Norwalk IAHA antibody, whereas 95% possessed antibody to the human rotavirus.
We have used an antigen solid-phase enzyme-linked immunosorbent assay (SP-ELISA) and an IgM antibody capture ELISA (MACELISA) for detecting IgM antibodies to human polyomavirus BK (BKV). These tests were compared with the standard hemagglutination inhibition test (HAI) of IgM serum fractions following sucrose density gradient fractionation. The SP- and MACELISA were not influenced by concomitant BKV-IgG, but high levels of both BKV-IgG and rheumatoid factor could cause false positive results by SPELISA, but not by MACELISA. The MACELISA gave much higher positive to negative ratios than the SPELISA. The sensitivity and specificity of the two tests were high compared to the IgM-HAI method. The sera could be tested in a single dilution (1:160), and thus the ELISA-tests are useful for testing large numbers of sera.
A technique that is based on absorption of sera with streptococcal cells and hemagglutination inhibition (HI) was evaluated for its feasibility for serologic diagnosis of recent rubella. The mixture of AR1 and AW43 cells removes IgG and IgA from the kaolin-treated sera, leaving IgM and a trace of IgA, probably oligomeric IgA. Consequently, after the absorption with streptococci, the HI antibodies are detectable exclusively in the early sera of patients with rubella. The streptococci (AR1 and AW43) have several advantages as the absorbent over the staphylococcus (Cowan I) that has been used routinely.
Extracellular components of plant pathogenic bacteria were obtained from their culture medium as well as from the whole cells by using NaCl 1 M, pH 6.0; 20% sucrose dissolved in 0.03 M Tris buffer, pH 8.0; or 0.05 M Na2EDTA. All the extracts from Erwinia carotovora subsp. carotovora, Xanthomonas campestris pv. campestris, Pseudomonas syringae pv. phaseolicola, Xanthomonas campestris pv. phaseoli, Pseudomonas solanacearum, and Erwinia carotovora subsp. atroseptica, were assayed for hemagglutinating activity on sheep, rabbit and chicken red blood cells (RBCs). The only active extracts were those obtained by NaCl treatment. They agglutinated sheep and rabbit erythrocytes. Extracts from E. carotovora subsp. atroseptica gave rise to the high agglutination titer on rabbit RBCs. These extracts had the lowest polysaccharide/protein ratio. E. carotovora subsp. carotovora extracts showed only a low titer (18.5 units). The agglutinating activity present in NaCl extracts of the bacteria tested was inhibited by different carbohydrates to various extent. Extracts from E. carotovora subsp. atroseptica appeared to be the most sensitive ones while those of E. carotovora subsp. carotovora least sensitive to the presence of sugar. It is suggested that hemagglutinins observed in plant pathogenic bacteria and those in plant host are similar and that both may, in some way, be involved in the plant-parasite relationship.
Kasba virus grown in BHK21-WI2 cells was tested for hemagglutination (HA) with erythrocytes of a variety of species at 4 degrees C, 25 degrees C and 37 degrees C. HA was observed at all temperatures with cattle, sheep and goat but not with swine, chicken, and goose erythrocytes. The HA was dependent on not only the NaCl concentration but also the pH of the diluent. The HA titer significantly improved by increasing the NaCl molarity to 0.6 M and standardizing pH to 7.5. The HA titer was 16- or 32-fold higher in 0.4 M or 0.6 M solution than in 0.2 M solution of not only NaCl but also several other salts. The HA reaction was inhibited by specific antibody.
Effects of methoprene and 20-hydroxyecdysone on the development and hemagglutination activity (HA) were studied in both sexes of two members of the Culex pipiens complex-anautogenous C. p. quinquefasciatus and autogenous C. p. molestus. Juvenile hormone analogue (methoprene) and 20-hydroxyecdysone caused developmental changes in both mosquito strains. High larval mortality, prolongation of intermolt period in each larval instar and in the pupal stage, and morphological changes in the larval-pupal and pupal-adult transformations were also observed. Developmental changes were accompanied with some differences in the HA. HA was found in both sexes of both experimental mosquito strains. The juvenile hormone analogue used in the larval stage caused significant decrease of HA in the gut of adults of both sexes. On the other hand, 20-hydroxyecdysone decreased HA only in the female gut. Results obtained indicate that HA depends on the sex, the studied organ, and the level of hormones.
When hemagglutinin (HA) of fowl plague virus (FPV) was expressed in CV-1 cells by a simian virus 40 vector, hemadsorption was barely detectable, although HA was exposed at the cell surface. However, treatment of HA-expressing cells with Vibrio cholerae neuraminidase (VCNA) resulted in extensive hemadsorption. VCNA treatment enhanced the electrophoretic mobility of the HA1 subunit of HA, indicating the removal of sialic acid. When two oligosaccharides in the vicinity of the receptor binding site of FPV HA were deleted by site-specific mutagenesis, VCNA treatment was not required for hemadsorption. Mutants which retained one of these oligosaccharides and mutants in which oligosaccharides not adjacent to the receptor binding site were deleted needed VCNA treatment to show hemadsorption. VCNA treatment also enhanced hemadsorption of vector-expressed HA of the WSN strain, which had a complex-type oligosaccharide in the vicinity of the receptor binding site, but had no effect on hemadsorption of Hong Kong type HA, which has a high-mannose type oligosaccharide adjacent to the receptor binding site. These results indicate that sialic acid on oligosaccharides near the receptor binding site interferes with hemadsorption. Thus, the neuraminidase is essential for FPV HA to show hemagglutinating activity.
The S protein of hemagglutinating encephalomyelitis virus is shown to be a hemagglutinin requiring N-acetyl-9-O-acetylneuraminic acid as a receptor determinant on the surface of erythrocytes. The ability of bovine coronavirus to recognize 9-O-acetylated sialic acid was used to establish a binding assay for the detection of glycoproteins containing this type of sugar. The assay is very fast, because it uses the acetylesterase of the viral HE protein to localize bound virus.