Specificity of anti-human IgG antibodies in antiglobulin (Coombs) sera.

The agglutination test with latex particles coated with aggregated human IgG was introduced into the evaluation of Coombs serum as an additional test for anti-IgG antibody activity. In Coombs sera prepared by the conventional immunization method employing Freund's adjuvant, latex agglutination titers were found much lower than those of anti-D-coated red cell agglutination. On the other hand, in sera prepared by other immunization methods, such as the one according to Haynes and Chaplin (1971), anti-IgG antibody response was readily observed by IgG-coated latex agglutination. Specificity of anti-IgG antibodies in the latter sera seems to be predominantly directed to aggregated human IgG.     

Serological investigation of Theileria sergenti using latex agglutination test in South Korea

Theileria sergenti causes persistent theileriosis in cattle, characterized by fever and chronic anemia. Theileriosis causes losses in feed efficiency and growth retardation through cycling infections in endemic areas. Among several major proteins of T. sergenti merozoites, the surface protein p33 is reported to be the most immunogenic. In this study, we investigated the use of p33 as a diagnostic antigen in a latex agglutination test to monitor antibodies against T. sergenti. When compared with TaqMan polymerase chain reaction, the sensitivity and specificity of the latex agglutination test were 86.5 and 92.5%, respectively. An epidemiological survey using the latex agglutination test was conducted with 1,046 sera collected from 4 slaughterhouses and 2 individual pasture farms throughout South Korea; 27.3% of samples were seropositive, depending on the areas in which the cattle were raised. This study indicated that the latex agglutination test could be used as a convenient tool for epidemiological monitoring of T. sergenti infections in the field.     

Commercially available histoplasmin sensitized latex particles in an agglutination test for histoplasmosis

Summary A commercial preparation of histoplasmin sensitized latex particles was tested in an agglutination test with sera from 50 culturally confirmed cases of histoplasmosis in varying stages of the infection. The reactions were superior to those obtained with collodion agglutination and complement fixation tests in which the antigen histoplasmin was also used. The latex agglutination test with the commercially available antigen is easy to do, and can be done in any laboratory equipped to carry out agglutination tests with the common bacterial antigens. It warrants more extensive trial in the general hospital laboratory as a screening test for histoplasmosis, especially the primary, pulmonary type.     

Evidence for Interference by Immune Complexes in the Serodiagnosis of Cryptococcosis

The latex agglutination test was used to compare cryptococcal antigen titers before and after protease treatment in 19 patients with pulmonary cryptococcosis. Antigen was detected by the LA test in 13 of 33 serum samples before protease treatment, and in an additional 13 samples following treatment. Of 26 antigen-positive serum samples, 22 (84.6%) showed an increased antigen titer after protease treatment. Using the cell slide agglutination test, antibody was detected in 3 of 19 cases. In one of these 3, antigen could only be detected after protease treatment. Soluble immune complex was prepared in vitro using anti-C. neoformans rabbit antiserum and polysaccharide antigen of C. neoformans, and the effects of immune complexes on the LA test were examined. In this experimental model, soluble immune complexes seemed to be observed in antibody excess region, because the antigen titers were increased after the protease treatment. We concluded that C. neoformans capsular polysaccharide antigen and anti-C. neoformans antibody formed soluble immune complexes in patients' sera, which interfered with antigen detection by the latex agglutination test without protease treatment.     

Exposure of platelet binding sites in von Willebrand factor by adsorption onto polystyrene latex particles

Von Willebrand factor molecules are flexible linear polymers composed of repeating protomeric polypeptide subunits. In the process of primary hemostasis, von Willebrand factor promotes platelet adhesion and platelet plug formation at the site of vascular injury. This biologic activity is apparently related to the multimeric size of von Willebrand factor. We simulated von Willebrand factor binding to the subendothelial surface by adsorbing purified human von Willebrand factor onto polystyrene latex particles of two different diameters, i.e., 0.312 μm and 2.02 μm. The rate and extent of ¹²⁵I-labeled von Willebrand factor binding to polystyrene was similar with both size classes of latex particles. The von Willebrand factor-coated latex beads of 2.02 μm diameter, in contrast to the smaller size, induced rapid agglutination of formalin-fixed human platelets in the absence of any other aggregating agent. Von Willebrand factor was also adsorbed from human plasma onto latex particles coated with anti-von Willebrand factor antibodies. Again, only the large beads, carrying the von Willebrand factor-antibody complex, induced agglutination of fixed platelets. Shear stress promoted the rate of von Willebrand factor adsorption to latex particles. Our results suggest that adsorption to surface exposes binding sites in human von Willebrand factor for platelets.     

Serological reactions associated with the clumping factor of Staphylococcus aureus

Rotter, Joan (University of Oklahoma Medical Center, Oklahoma City), and Florene C. Kelly. Serological reactions associated with the clumping factor of Staphylococcus aureus. J. Bacteriol. 91:588-594. 1966.-Evidence that the substance which causes staphylococci to clump in the presence of fibrinogen (clumping factor) is antigenically similar in strains which are serologically diverse according to agglutination reactions has been obtained from fibrinogen-cell clumping inhibition tests. Antisera for clumping factor (CF)-positive strains inhibited the clumping reaction of all strains tested. After adsorption with homologous cells or with cells of other CF-positive strains, the antisera no longer inhibited clumping. When antisera were adsorbed with trypsin-treated, CF-positive cells, or with cells of CF-negative mutants, the ability to inhibit the clumping reaction persisted. Antibody to CF activity was not associated with coagulase. Latex coated with extracts derived from the cells of five CF-positive and six CF-negative strains was, in each instance, agglutinated by sera from rabbits immunized with CF-positive cells. After adsorption with trypsinized, CF-positive cells, antisera still agglutinated latex which had been treated with the CF-positive extracts, but not with the CF-negative extracts. Similar results were obtained after antisera were adsorbed with the cells of CF-negative mutants. Cell agglutination titers of sera from rabbits immunized with CF-negative staphylococci were significantly lower than those produced in response to CF-positive cells, regardless of their coagulase activity. If the CF-inhibiting antibody also functions as an agglutinin, it apparently is not solely responsible for this difference.     

A trial application of the latex test for evaluation of antibody level in rabbit immune sera]

A latex test was elaborated which served for evaluation of quality of rabbit immune sera for antigen 0 of selected Gram-negative bacteria. Sensitivity and specificity of this test in comparison with passive hemagglutination and immunoenzymatic DOT-ELISA reactions was evaluated. These studies were performed on immune sera for antigen O of Salmonella groups B, C1, C2, D and E, Yersinia pseudotuberculosis and in antigen preparations for above listed microorganisms both in homologous and heterologous systems. It was found that sensitivity of the latex test is 9 to 160 times lower than that of passive hemagglutination and 7 to 307 lower than for DOT-ELISA. Sensitivity of the latex test and passive hemagglutination reaction was evaluated on the basis of results of cross reaction between studied antigens and unabsorbed rabbit sera, establishing so called sensitivity indexes, which were informing how many times heterologous titer is lower than homologous titer. So evaluated sensitivity of the latex test was close to sensitivity of the passive hemagglutination reaction. It was found that slide latex test is characterized by satisfactory sensitivity and good sensitiveness and may be used for evaluation of antibody level 0 antigens of Salmonella and Yersinia. The value of this test is characterized by high repeatability of results, as well as low work and time-consuming.     

The antigen-antibody reaction and its inhibition

The sera of intact and immune animals contain factors capable of inhibiting the interaction of antibodies with the homologous antigen. These agglutination inhibiting (AI) factors not only block the antigen, thus competing with antibodies, but may induce the dissociation of the antigen-antibody complex. Heating of antisera at 60-63 degrees C leads to an increase in the activity of AI factors. The phenomenon of "prozone" in the agglutination test appears to be explained by the presence of AI factors in sera.     

Antibodies against interspecies erythrokaryocyte antigen in patients with partial red cell aplasia of the bone marrow

Immunofluorescense and cytotoxicity test in vitro were used to demonstrate specific antibodies in sera of 11 out of 19 patients with partial red cell aplasia (PRCA). The antibodies reacted with erythroblast cells from embryos and adult men, with bone marrow cells from a female patient suffering from acute erythroleukemia, with erythrokaryocytes of mouse embryos and cells of Rauscher's viral erythroleukemia. The results of cross adsorption and blockade of the immunofluorescent reaction of the sera of PRCA patients with antibodies against mouse erythroblast antigen bearing an interspecies determinant suggest that in the pathogenesis of PRCA there takes part an autoimmune reaction against specific interspecies antigen to erythrokaryocytes. This antigen is apparently similar to antigen against mouse erythroblast cells.     

Immunological properties of an L-form of a group A beta hemolytic streptococcus

Antisera were prepared by injecting several groups of rabbits with antigen preparations of the parent streptococcus and its derived L-form. Serum was obtained from each rabbit by cardiac puncture prior to and at intervals after injection of the organisms. To measure the immune response of all animals a modified latex agglutination test was used. This test was found to be specific and easily reproducible. Animals immunized intravenously with the heat-killed streptococci and those immunized intramuscularly with the L-form produced the highest level of antibodies against their respective antigen preparations. Lancefield extracts of the streptococcus reacted with the streptococcal antisera but not with antisera to the L-form. In the antisera to the L-form we found specific antibodies which could not be adsorbed with the parent streptococcus.A portion of this study formed part of a thesis submitted by Sister M. B. Muellenberg to the University of South Dakota, Vermillion, S.D. in partial fulfillment for the requirement for the degree of Master of Arts.     

Trypanosoma lewisi: Avidity and adsorbability of ablastin,the rat antibody inhibiting parasite reproduction

The relation of naturally acquired host IgG in the surface coat of bloodstream forms of Trypanosoma lewisi to ablastin was studied to determine whether, contrary to a long-held conclusion, the antibody is avid and adsorbable. It was found by immunofluorescence and agglutination tests with monospecific antisera to rat IgG that bloodstream forms collected from immunosuppressed hosts, in contrast to those from immunocompetent hosts, have little or no detectable surface IgO. Specificity of adsorption was also demonstrated in other immunofluorescence experiments in which bloodstream forms from immunosuppressed hosts adsorbed IgG from immune serum with ablastic activity only (previously adsorbed with trypanosomes from immunocompetent hosts to remove the trypanocidal antibodies), but did not adsorb IgG from normal rat serum. To determine whether this specific adsorption of IgG by the parasite could be correlated with a reduction in ablastic activity, immune sera were adsorbed with bloodstream forms from immunosuppressed hosts at packed cell/serum ratios of either 1.2 or 2.0, and the adsorbed sera were then tested for ablastic activity in vitro. With both cell/serum ratios, ablastic activity was reduced by 50%. In comparison, similar adsorptions of immune sera with trypanosomes from immunocompetent hosts resulted in reductions of ablastic activity of only about 9 and 27% with the low and high cell/serum ratios, respectively. It is concluded that the failure to effect significant adsorption of ablastin in earlier studies resulted from the use of ablastinsensitized trypanosomes from immunocompetent hosts.     

Trypsinized Human Group O Erythrocytes as an Alternative Hemagglutinating Agent for Japanese Encephalitis Virus

Trypsinized human group O erythrocytes were found to be a suitable alternative to gander cells in hemagglutination (HA) and hemagglutination inhibition (HAI) tests for Japanese encephalitis (JE) virus. In the HAI test, no cross-reactions against JE virus were observed with immune sera containing antibody to taxonomically related or unrelated viruses, with mouse brain antigen, or with nonantibody serum inhibitors; specific antibody rise could be detected in an immunized rabbit. Gander and trypsinized human group O cells gave comparable titers in the HAI test, but the latter were preferable since (i) they required less challenging HA antigen, being more sensitive to agglutination by JE virus, and (ii) all human and some animal sera investigated were devoid of natural agglutinins for these cells, thereby eliminating or reducing the need for prior adsorption with packed cells.     

A rabies agglutination test (RAT) for rabies antibody detection

An agglutination test has been developed for the detection of rabies antibodies after human vaccination. The rabies agglutination test (RAT) is based on the capability of specific antibody to agglutinate sensitized polystyrene (or latex) beads. In the RAT, latex beads were coated, in a first step, with inactivated and purified rabies virus (PV strain adapted and propagated on BHK-21 cells) and, in a second step, with bovine serum albumin. Negative control beads were coated with bovine serum albumin (BSA) only. To test for human antibody, several microliters of serum were mixed on a glass slide with an equal volume of virus-sensitized beads and the mixture was gently agitated. After a few minutes, agglutination was visible with sera which had been characterized as positive by the virus neutralization antibody (VNAb) technique. No agglutination was observed with negative sera tested with virus-coated beads or with positive sera tested with BSA-coated beads. Virus-sensitized beads were agglutinated when the virus neutralizing antibody titres were equal to or greater than 2.5 international units per ml (IU/ml) in human sera. The concordance between the RAT results and VNAb titres was about 97% when 2.5 IU/ml was taken as the cut off value for determining the positive sera with the VNAb technique. The possibility that clinicians might use the RAT as a simple means to determine sero-conversion at the end of the post-exposure treatment of patients is discussed.     

Lipopolysaccharide O-antigen antibody-based detection of the fish pathogen Flavobacterium psychrophilum

Several yellow-pigmented species within the family Flavobacteriaceae are commonly associated with diseases in fish and are difficult to speciate due to their fastidious, slow-growing nature and cross-reactive antigens. Here we report the development of specific, antibody-diagnostic tests for Flavobacterium psychrophilum, the aetiological agent of rainbow trout fry syndrome and bacterial cold water disease. A unique antigen from F. psychrophilum, the lipopolysaccharide (LPS) O-polysaccharide (O-PS), formed the basis for the antibody test. LPS O-PS was purified and conjugated to keyhole limpet haemocyanin and bovine serum albumin for the generation of rabbit immune sera and the development of antibody-based diagnostic tests. Rabbit polyclonal anti-O-PS serum was highly specific for F. psychrophilum, without the need for prior cross-absorption with related bacteria and was the basis of an effective ELISA diagnostic test. Antibodies were purified from rabbit anti-O-PS serum and adsorbed onto coloured latex beads for the development of a specific, bead agglutination assay for F. psychrophilum.     

Detection of Chlamydia species-specific serum antibodies by prior adsorption of common genus-specific antibodies

To establish a method for the detection of Chlamydia species-specific antibodies to the three species of Chlamydia responsible for human disease, the author attempted to remove Chlamydia genus-specific antibodies by prior adsorption with heterologous Chlamydia antigen. The effects of adsorption with heterologous antigen were investigated by the microplate immunofluorescence antibody technique. The Chlamydia genus-specific antibodies in immune animal sera were significantly reduced by prior adsorption with heterologous Chlamydia antigen. Chlamydia pecorum which does not infect humans was found to be useful for the adsorption. A preliminary test using Chlamydia trachomatis-infected human sera showed that this adsorption method with C. pecorum is applicable to the serodiagnosis of human Chlamydia infections.     

Value and Interpretation of Serological Tests for the Diagnosis of Cryptococcosis

MAXIMAL SEROLOGICAL DIAGNOSIS OF CRYPTOCOCCOSIS MAY BE ACCOMPLISHED THROUGH THE CONCURRENT USE OF THREE TESTS: the latex agglutination (LA) test for cryptococcal antigen, and the indirect fluorescent antibody (IFA) and tube agglutination (TA) tests for Cryptococcus neoformans antibodies. These tests were applied to 141 serum and cerebral spinal fluid specimens from 66 culturally proven cases of cryptococcosis and to 42 sera from normal subjects and from patients with other systemic mycotic diseases. The LA test was sensitive and completely specific; of the sera from proven cases, 55% were positive. With the TA test, 37% of the specimens were positive and the test was highly specific. With the IFA test, 38% of the specimens were positive and the test appears to be the least specific of the three. Cross-reactions were most evident with blastomycosis and histoplasmosis case sera. When the three tests were used concurrently, 87% of the cryptococcosis case specimens were positive and permitted a presumptive diagnosis of C. neoformans infections in 61 (92%) of the 66 patients whose specimens were examined.     

Comparative evaluation of whole blood D-dimer test to plasma D-dimer test for diagnosis of disseminated intravascular coagulation

Three rapid D-dimer test methods were compared for the diagnosis of acute disseminated intravascular coagulation (DIC). These were (a) SimpliRED, an autologous red cell agglutination assay. (b) DIMERTEST latex agglutination assay, containing monoclonal antibody DD-3B6/22(6), and (c) D-DI latex agglutination assay containing mouse anti-human D-dimer monoclonal antibodies. The D-DI latex method having higher sensitivity (100%) and specificity (81%) in clinically acute DIC was postulated as the gold standard and compared with the other two methods. The results suggest that D-DI latex agglutination assay containing mouse anti-human D-Dimer monoclonal antibodies are the better assay methods amongst all the three kits analyzed. It is advisable to look for the nature of the antibody used to coat the latex particles in plasma based kits. In emergency setting RBC kits may be of some use as rapid diagnosis is advantageous.     

Determination of antibodies to diphtheria and tetanus toxoid by latex agglutination technique

A micromethod of latex agglutination was worked out for determination of antibodies to diphtheria and tetanus toxoids. It is suitable for pediatrics since a minimum amount of serum (0.05 ml.) is required, which can be used without being inactivated or absorbed. Particles of polystyrene latex of Czechoslovak production were used as antigen carriers thus enabling better standardization of the technique. The particles are well defined both in physical and chemical terms and replace red blood cells in passive hemagglutination. The method was compared with passive hemagglutination in a dynamic study of children inoculated at monthly intervals with the trivalent vaccine Alditepera.     

Serological Activity of Staphylococcal Polysaccharide

The polysaccharide from cell walls of coagulase-positive staphylococci coated both latex particles and tanned red cells for agglutination by human sera and by specific staphylococcal antisera. Treatment with trypsin or autoclaving destroyed the capacity of polysaccharide to coat particles but did not affect precipitation of antibody. Periodic acid destroyed both properties. The teichoic acid portion of the staphylococcal polysaccharide displayed precipitin activity similar to polysaccharide, but it did not coat either latex particles or tanned red cells. Teichoic acid did, however, inhibit specific agglutination of polysaccharide-coated particles or cells.