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PubMed1979

Improved indirect hemagglutination test for cytomegalovirus using human O erythrocytes in lysine.

A modified indirect hemagglutination test for cytomegalovirus antibodies is described in which glutaraldehyde-fixed human O cells, rather than sheep cells, are used. Nonspecific hemagglutination was reduced by use of the optimal tannic acid concentration for each cell batch and the addition of 0.1 M lysine to the phosphate-buffered saline in which the fixed, tanned, sensitized cells were resuspended for use in the test. Three of 349 sera showed nonspecific hemagglutination by this technique. Antigen made by freezing phosphate-buffered saline (pH 7.2) over an infected monolayer can be used at dilutions of 1:8 to 1:15. Fixed, tanned, sensitized cells ready for use in the test can be stored in liquid nitrogen for up to 8 months. Use of cryoprotectants and washing after thawing is unnecessary. Simplification of the assay permits one person to screen 300 sera in 1 day or to determine the immune status of a potential donor or recipient in 45 min after the test is set up. The modified assay compares favorably in sensitivity with the complement fixation test and with previously described methods for performing the indirect hemagglutination test.

ABO Blood-Group System↗

PubMed1977

Use of sodium polyanetholesulfonate-CaCl2 for removal of serum nonspecific inhibitors of rubella hemagglutination: comparison with other polyanion-divalent cation combinations.

By using trypsin-treated human type O cells as indicators, we compared the abilities of four polyanion-divalent cation combinations (heparin-MnCl(2); high-and low-molecular-weight dextran sulfate-CaCl(2); and sodium polyanetholesulfonate [SPS]-CaCl(2)) for removal of serum non-immunoglobulin (lipoprotein) inhibitors of rubella hemagglutination. The combination of SPS-CaCl(2) was found to be the most effective, precipitating completely the pre-beta and beta-lipoproteins and reducing the alpha-lipoprotein levels by more than 50%. Hemagglutination patterns after this treatment were clear and stable, and, when normal sera were tested, hemagglutination-inhibition (HI) titers were comparable to those obtained after standard heparin-MnCl(2) treatment. High-molecular-weight dextran sulfate-CaCl(2) removed serum lipoproteins almost as effectively as SPS-CaCl(2). However, problems of nonspecific agglutination and the heavy hemagglutination patterns resulting made this combination unacceptable for routine purposes. Neither low-molecular-weight dextran sulfate-CaCl(2) nor heparin-MnCl(2) removed the pre-beta lipoproteins completely, and occasionally traces of beta-lipoprotein also remained after treatment. The presence of pre-beta lipoproteins in normal sera after treatment may be of no consequence in the HI test since we have found that the very-low-density lipoprotein fractions obtained by ultracentrifugal methods from normal sera (those corresponding to the pre-beta fractions obtained by electrophoresis) had no HI activity. However, very-low-density lipoprotein fractions from all hyperlipemic sera tested had HI activity (titers ranging from 1:16 to 1:1,024) which, in the majority of cases, was not eliminated after heparin-MnCl(2) treatment. In every case, treatment with SPS-CaCl(2) removed this nonspecific activity completely. Since hyperlipemic sera may occasionally be encountered in routine rubella HI antibody testing, we recommend the use of SPS-CaCl(2) rather than heparin-MnCl(2) for pretreatment of sera.

Antibodies, Viral↗

PubMed1978

Quantitation of antibodies to varicella-zoster virus by immune adherence hemagglutination.

Immune adherence hemagglutination was compared with the complement fixation test as a means of measuring antibodies to varicella-zoster virus. Analysis of acute- and convalescent-phase sera from patients infected with varicella-zoster or with herpes simplex virus showed the immune adherence hemagglutination test to be more sensitive than the complement fixation test, and greater cross-reactivity between the two viruses appeared to be associated with the increased sensitivity. The two assay methods were used to measure antibodies to varicella-zoster virus in 265 sera obtained from patients of different ages as well as sera from 26 patients with leukemia. There were 35 cases where antibodies were detected by immune adherence hemagglutination but not by complement fixation, whereas in five cases the converse was found. Our findings support the contention that immune adherence hemagglutination is the method of choice for detecting antibodies to varicella-zoster virus.

Acute Disease↗

PubMed1975

Fibrinogen-fibrin degradation products: hemagglutination inhibition immunoassay in plasma.

Immunoassay for fibrinogen and/or fibrin degradation products (FDP) is generally in the clot and hence assay of serum may not reveal the true concentration of FDP in blood. We have developed a hemagglutination inhibition immunoassay for FDP in human plasma. D fragment appears to possess an antigenic determinant, called D-neoantigen, not found in native fibrinogen. Rabbit antiserum produced against D fragment was absorbed with immunosorbent columns coupled with fibrinogen and normal human serum, respectively, so that it contained only those antibodies directed against the neoantigenic determinant of D fragment. In this immunoassay, sheep erythrocytes (SRBC) were stabilized with glutaraldehyde and subsequently sensitized with D fragment by means of tannic acid. Hemagglutination of absorbed anti-D-neoantigen serum against SRBC sensitized with D fragment was titered to be 1:256. The hemagglutination was inhibited by D fragment but not by fibrinogen; the sensitivity of detecting D fragment was 8 mug/ml. Human plasma from normal subjects did not inhibit. This appears to be the first report of a hemagglutination inhibition immunoassay for FDP in plasma.

Animals↗

PubMedDate not supplied

[The detection of rheumatic factors by means of a new hemagglutination drop test (Cellognost-RF)].

A comparative evaluation was made in 510 sera to study the specificity and sensitivity of 5 different methods for the determination of rheumatoid factors. Special attention was given to the hemagglutination drop tests which are commercially available (Cellognost-RF and Rheumaton). Both hemagglutination slide tests, which are easy to perform and easy to read, were shown to be practically equivalent, and in comparison to the other methods, also showed a good agreement. This is as well applicable to rheumatoid arthritis as to non-inflammatory processes and internal diseases. In practice, Cellognost-RF and analogous hemagglutination slide tests are certainly good screening tests for the rheumatic factors. In combination with the also simple latex-slidetest, the detection of rheumatoid factors in rheumatoid arthritis is somewhat better, although not much. When both reactions are negative, it is not necessary to do any additional tests for the demonstration of rheumatoid factors. Should one or both, however, be positive, the titre determination using the hemagglutination reaction of Waaler-Rose or latex fixation test should be made if possible. In our investigations it was also in principle possible to do these titre determinations with Cellognost-RF.

Adolescent↗

PubMed1978

Leptotrichia buccalis hemagglutination in cell binding and salivary inhibition studies.

The characteristic hemagglutination (HA) of Leptotrichia buccalis was used for measuring its attachment to various human cells and for determining if saliva contained hemagglutination inhibition (HI) factors. The microbial strain utilized displayed the characteristic EM morphology of L. buccalis. Sonicated preparations of the organism were tested for HA activity before and after adsorption with human cells. Buccal epithelial cells, red blood cells (RBC), HeLa and embryonic kidney cells all bound the HA fragments of the organisms. The bacterial fragments on the cells could be observed by fluorescent antibody testing. The fragments were released from the cells used for adsorption with chelators and upon addition of CaCl2 the HA activity returned. Whole saliva displayed hemagglutination inhibition activity in a manner suggesting a binding site interaction. The similarity of the HA activity of F. nucleatum is discussed as are the relationships of cell binding to colonization of the organisms and immunopathology to host cells.

Adsorption↗

PubMed1976

Hemagglutination by equine infectious anemia virus.

Equine infectious anemia (EIA) virus which was propagated on an equine dermal cell line agglutinated guinea pig erythrocytes. Viral fluids containing about 10(7.5) mean tissue culture infective doses/ml showed hemagglutinating (HA) titers ranging from 16 to 32 units/0.05 ml. Results of cesium chloride equilibrium density gradient centrifugation revealed that the hemagglutinin was inseparable from the virus particles. The hemagglutination reaction persisted over a wide range of temperature and pH, and the absence of divalent cations did not decrease its activity. The HA activity was stable at 4 degrees C but not at 56 degreesC. The activity was destroyed by virus-disrupting lipid solvents and moderately sensitive to a proteolytic enzyme. Neuraminidase enhanced HA activity slightly. Phospholipase C had no effect on HA titer, although it completely inactivated infectivity. It was relatively stable to ultraviolet irradiation. Thus, the hemagglutinin appears to be closely associated with virus particles, and its activity is dependent on the presence of its lipids and proteins. Hemagglutination was inhibited by sera from horses infected with EIA virus. Hemagglutinin receptors on the erythrocytes were inactivated by a proteolytic enzyme and formaldehyde but were not influenced by neuraminidase, sodium deoxycholate, or KIO4.

Animals↗

PubMed1978

Detection of early "significant" antibody responses in paired sera by the use of an automated bovine parainfluenza-3 hemagglutination-inhibition system.

An automated bovine parainfluenza-3 hemagglutination-inhibition system was used to detect small increases (15--20%) in specific antibody titer between paired bovine sera collected three days apart. Automated and manual parainfluenza-3 hemagglutination-inhibition test procedures were compared as methods for the serologic diagnosis of parainfluenza-3 infection. Fifteen daily serum specimens from each of three animals were analyzed as various possible manual-test and automated-test serum pairs. With the sera from two of the animals, considered to be parainfluenza-3 hemagglutination-inhibition-positive, the manual microtiter test procedure made 18 (69%) false negative diagnoses when using the standard manual-test criteria for "evidence of infection." When a 20% titer increase between three-day serum pairs was considered "serologically significant," the automated system made three (17%) false-negative diagnoses. At a 15% "serologically significant" increase, no false-negative diagnosis was made. The utilization of automated serology for the routine diagnosis of disease should permit treatment and control measures to be initiated early, thereby reducing losses due to infectious disease.

Animals↗

PubMed1977

Comparison of the enzyme-linked immunosorbent assay and the indirect hemagglutination test for detection of antibody to cytomegalovirus.

Thirty samples of serum were tested for antibody to cytomegalovirus by enzyme-linked immunosorbent assay (ELISA) and indirect hemagglutination (IHA). The two tests were in extremely close agreement. Of the 30 sera evaluated by ELISA, 16 were considered to be positive and 14 were considered negative. In all 16 positive samples, the titers by ELISA were at least twofold to 10-fold higher than the indirect hemagglutination titers. Two of the 14 sera evaluated as negative by the ELISA test had low indirect hemagglutination titers (1:8 and 1:4). They were not detectable at the initial dilution (1:50) of the ELISA test. These two sera might have been classified as positive if the initial dilution in ELISA had been lower than 1:50.

Animals↗

PubMed1977

Restitution of hemagglutinating activity to spikeless particles of HVJ (Sendai virus) by glycoprotein components of Newcastle disease virus.

Spikeless particles of HVJ (Sendai virus) lacking in hemagglutinating (HA) activity were obtained by enzymatic digestion of virions with trypsin followed by centrifugation through a sucrose gradient. When they were mixed with glycoprotein components of Newcastle disease virus (NDV) obtained by treatment of purified virions with deoxycholate (DOC), the mixture showed hemagglutination reaction, which was inhibited by anti-NDV serum, but not by anti-HVJ serum. Sedimentation profile of the HA active agents was then examined by centrifugation of the mixture of spikeless particles of HVJ (labeled with 3H-uridine) and glycoproteins of NDV (labeled with 14C-amino acid mixture). The results showed that the peak of HA activity had both of the radioactivities, and that the sedimentation rate of the HA was faster than that of spikeless HVJ but slower than that of intact HVJ. Electron micrographs of such HA active structures showed that they were morphologically closely similar to intact virion of HVJ, although they had neither hemolytic activity nor infectivity. The mixture of spikeless HVJ and glycoproteins of HVJ or NDV which were removed from virions by proteolytic enzymes, on the other hand, did not show any detectable hemagglutinating activity.

Deoxycholic Acid↗

PubMed1978

Heterophil antigen in bovine sera detectable by immune adherence hemagglutination with infectious mononucleosis sera.

Bovine sera for tissue culture use were shown to contain heterophil antigen of the Paul-Bunnell-Davidsohn type by immune adherence hemagglutination tests with sera from patients with infectious mononucleosis. The antibody titers observed were comparable to those determined by two other methods; i.e., the immune adherence hemagglutination test was found to be as specific as the differential absorption test with horse erythrocytes or the ox cell hemolysis assay,but it appeared to be the most sensitive of the three procedures. All 16 individual or pooled bovine serum samples showed Paul-Bunnell-Davidsohn antigen at concentrations varying over a fourfold range. The potential usefulness of the immune adherence hemagglutination test for various Paul-Bunnell-Davidsohn antigen-related problems and implications of the observations are discussed.

Animals↗

PubMed1978

Association of hemolysin production, hemagglutination of human erythrocytes, and virulence for chicken embryos of extraintestinal Escherichia coli isolates.

One hundred forty-two strains of Escherichia coli isolated from extraintestinal infections were examined for colicin V (ColV) and hemolysin (Hly) production. For comparison, 20 strains isolated from the feces of normal individuals and 12 enteropathogenic strains of E. coli were tested for these properties. Thirty-five to 59% of extraintestinal isolates were Hly+, but only one fecal strain was Hly+. Colicin V biosynthesis was found for 12% of blood culture isolates, 7% of urine culture isolates and 16% of the strains from other extraintestinal infections. None of the fecal isolates was ColV+. Selected strains were tested for virulence in 13-day-old chicken embryos; these same strains were tested for their ability to hemagglutinate chicken or human erythrocytes. Of 22 extraintestinal isolates, 13 (59%) killed greater than or equal to 60% of the embryos within 72 h. Only one of eight normal fecal isolates and two of three enteropathogenic strains tested were virulent. About 80% of the virulent strains were Hly+. The most striking finding, however, was the hemagglutination of human erythrocytes by virulent extraintestinal isolates. It seems possible that the hemagglutination property reflects a specific common adherence factor.

Animals↗

PubMed1979

Indirect hemagglutination test for detection of antibody to Rickettsia rickettsii in sera from humans and common laboratory animals.

Antibody production in humans and three species of laboratory animals infected with Rickettsia rickettsii was determined with the indirect hemagglutination test. Rabbits, guinea pigs, and mice were inoculated with R. rickettsii and bled at intervals. Antibody which agglutinated both fresh and glutaraldehyde-fixed sheep erythrocytes sensitized with antigen prepared either from purified rickettsiae or from infected yolk sacs was found in rabbit sera at all intervals tested (10 to 59 days postinfection). Antibody which agglutinated fresh but not glutaraldehyde-fixed erythrocytes sensitized with either of the above antigens was detected in guinea pig sera obtained 7, 14, and 28 days postinfection. Antibody was found in mice inoculated with 5.6 x 10(6) plaque-forming units of R. rickettsii but not in mice given 5.6 x 10(2) plaque-forming units. Peak indirect hemagglutination titers occurred in nonvaccinated human Rocky Mountain spotted fever patients about 3 weeks after onset of illness, and antibody was still detectable after 1 year. Both human immunoglobulin G and human immunoglobulin M antibodies agglutinated sensitized cells, but immunoglobulin M antibodies apparently were more efficient. The indirect hemagglutination test is useful for the titration of human, rabbit, guinea pig, and mouse antibodies when the appropriate erythrocytes are used.

Animals↗

PubMed1979

Analysis of Vibrio parahaemolyticus soluble antigens by employing passive hemagglutination.

The passive hemagglutination assay was explored as a sensitive test of immunological reactivity of endotoxin and other antigens prepared from selected strains of Vibrio parahaemolyticus. The tannic acid procedure for passive hemagglutination, commonly used with protein antigens, was the only procedure yielding good results with V. parahaemolyticus protein extracts, endotoxins, and related preparations. These results were probably due to the presence of large amounts of protein in the V. parahaemolyticus endotoxins as determined by earlier work referenced in the text. Glucose and galactose as possible antigenic determinants in the endotoxin of a Vibrio strain were tested by inhibition tests. Cross-reactions were observed between endotoxin preparations, but were low in hemagglutination, suggesting reactions of common generic antigens. The ability of V. parahaemolyticus endotoxins to stimulate production of antibodies was determined.

Antibodies, Bacterial↗

PubMed1979

Comparison of indirect hemagglutination and indirect immunofluorescence tests with microneutralization tests for detection of type-specific Herpesvirus hominis antibody.

Indirect hemagglutinating and immunofluorescent antibody responses to Herpesvirus hominis types 1 and 2 were compared to neutralizing antibody responses in infected humans from whom H. hominis type 1 or 2 was isolated. The indirect immunofluorescent antibody test was shown to be the most sensitive and specific for primary human infections. The sensitivity and specificity of the indirect hemagglutination and the immunofluorescent antibody tests were shown to be equal to that of the microneutralization test among patients who had primary or recurrent H. hominis type 2 infections. It is suggested that the indirect hemagglutination test is preferable for assaying large populations for previous infection with H. hominis type 2 because it is rapid, easier to perform, and more economical. The intermediate range of titer differences (deltat) between H. hominis types 1 and 2 previously reported to be due to infections with both viruses was shown to occur in all three tests among patients with primary infections with either virus.

Antibodies, Viral↗

PubMed1978

[Use of the indirect hemagglutination reaction for detecting antibodies to Pseudomonas pyocyanea in acute suppurative destructive pneumonia].

No less than 4-fold increase in the antibody titres to Pseudomonas aeruginosa during the infectious process served as laboratory confirmation of its participation in the infectious process. In 49 of 91 patients hemagglutining titre exceeded the diagnostic one (1:640). In 9 patients (chiefly in infants) hemagglutinin titre remained low, but there was a rise of antibody level to Pseudomonas aeruginosa during the disease. High hemagglutinin titres were noted in 12 patients on admission to the clinic, with reduction of the antibody titres at the late periods of the disease. Antibody titres remained unchanged during the disease in 15 patients. In 3 cases the indirect hemagglutination test was assesed as negative. In the rest of the patients hemagglutinin titres varied within the range of the diagnostic titre. Thus, the indirect hemagglutination test with erythrocytic diagnostic agent permitting to determine antibodies to Pseudomonas aeruginosa could be used at the clinic in combination with bacteriological and other investigations for establishing etiology of the destructive process.

Acute Disease↗

PubMed1976

[Sensitivity and specificity of the passive hemagglutination reaction applied to titration of herpes antibodies].

52 human sera were checked for herpes virus hominis type 1 antibodies by passive hemagglutination, complement fixation, neutralization tests and plaque reduction in presence of complement. A good correlation was observed among the results obtained by the passive hemagglutination test and the plaque reduction in presence of complement. The rapidity of the indirect hemagglutination test and the sensitivity of the reaction are in favour of its routine use.

Antibodies, Viral↗

PubMed1976

[Indirect hemagglutination test for detection of antibodies to cytomegalovirus in blood collected on blotting paper (author's transl)].

Indirect hemagglutination test for detection of antibodies to cytomegalovirus is highly sensitive and reproducible, if employed in well-defined conditions. Standardization of the various factors involved is necessary as well as their reciprocal equilibrium : sheep erythrocytes, antigen, dilution of tanin, buffers quality. The hemagglutination test can be performed on small volumes such as blood collection on blotting paper (PKU). Antibody titers were compared in the serum and the blood so collected in 104 subjects : the results were very similar and no "false negative" were found in any case. This way of collecting blood and hemagglutination are technical improvements in epidemiologic studies of cytomegalovirus infection. It can be hoped they will be adapted to other group herpes infections.

Animals↗