[Diagnosis of human brucellosis. Role of pH in the seroagglutination test and influence of pH on the agglutinating activity of IgM, IgG and IgA antibodies].
OBJECTIVE
To evaluate the role of pH in the seroagglutination test (SAT)and Rose Bengal (RB) test, and to determine the influence of pH on the agglutinating activity of IgM, IgG and IgA antibodies.


METHODS
The SAT was performed at pH 7.2 or pH 5.0 in standard microtiter-type polystyrene plates using Ring Test antigen or the Brucella suspension (BRUCAPT) provided in the Brucellacapt kits. Specific antibodies against native hapten were determined by radial immunodiffusion. Additionally, IgG, IgA and IgM fractions were separated from 8 sera by absorption chromatography and their agglutinating capacity was studied at pH 7.2 and 5.0.


RESULTS
We studied 72 sera from patients with clinical brucellosis taken at the time of hospitalization, 16 from persons in contact with infected animals, and 16 from healthy donors. SAT results at pH 5.0 correlated with those obtained with the Rose Bengal test. Four Rose Bengal-positive sera were found to be SAT-negative at pH 7.2 and SAT-positive at pH 5.0. SAT performed at pH 5.0 with BRUCAPT antigen yielded higher titers than tests performed at pH 7.2 or 5.0 with Ring Test antigen (p < 0.001), with highest titers in IDR-positive sera. Among the 8 IgG fractions, all but one agglutinated at pH 7.2, and in 4, IgG titers showed significant increases at pH 5.0. Three IgA fractions were SAT-negative at pH 7.2 and SAT-positive at pH 5.0; the other 5 agglutinated at both pH conditions and were DTT-sensitive. All IgA fractions but one were positive by Rose Bengal. Agglutinating activity of the IgM fraction was not affected by pH.


CONCLUSIONS
The SAT performed with the buffer and antigen suspension included in the Brucellacapt kit (pH 5.0) is highly useful for detecting agglutinating and non-agglutinating antibodies at pH 7.2.
Rubio Vallejo
Manuel
M
Departamento de Microbiología. Servicio de Microbiología. Clínica Universitaria. Universidad de Navarra. Pamplona. Spain.
del Pozo
José L
JL
Del Pozo León
José Luis
JL
Hernández-Molina
Juan Manuel
JM
Dorronsoro Ibero
Inés
I
Marrodán Ciordia
Teresa
T
Díaz García
Ramón
R
D000372:Agglutination Tests / Q000379:methods*; D000907:Antibodies, Bacterial / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D000942:Antigens, Bacterial / Q000276:immunology; D016470:Bacteremia / Q000175:diagnosis / Q000276:immunology / Q000382:microbiology; D017347:Brucella melitensis / Q000276:immunology*; D002006:Brucellosis / Q000175:diagnosis* / Q000276:immunology / Q000382:microbiology; D005591:Chemical Fractionation; D004396:Coloring Agents / Q000737:chemistry; D003378:Counterimmunoelectrophoresis; D004229:Dithiothreitol / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D005779:Immunodiffusion; D007070:Immunoglobulin A / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D007075:Immunoglobulin M / Q000276:immunology; D008070:Lipopolysaccharides / Q000276:immunology; D011933:Reagent Kits, Diagnostic; D011934:Reagent Strips; D012395:Rose Bengal / Q000737:chemistry
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D004396:Coloring Agents; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D008070:Lipopolysaccharides; D011933:Reagent Kits, Diagnostic; D011934:Reagent Strips; D012395:Rose Bengal; D004229:Dithiothreitol