A specific and ultrasensitive chemiluminescent sandwich ELISA test for the detection and quantitation of pneumolysin.
A chemiluminescent sandwich ELISA test has been developed for the detection and quantitation of pneumolysin. The test is based on a mouse monoclonal as the capture antibody and on rabbit polyclonal IgGs as detection antibodies, in combination with an anti-rabbit IgG alkaline phosphatase conjugate. The estimated detection limit of the purified recombinant toxin in phosphate-buffered saline with 0.05% Triton X-100 is around 5 pg ml(-1), with averaged intra- and inter-assay variation coefficients of 7% and 13.5%, respectively. The assay has been applied to the quantitation of pneumolysin in pneumococcal isolates, providing, for the first time, a direct measurement of the amount of the toxin produced by different strains, a variation has been found in their pneumolysin content. The test is highly specific as no other purified toxins or human pneumonia- or meningitis-associated bacteria yielded false-positive results. This specific and highly sensitive method could help in the diagnosis of human infections.
Cima-Cabal
M D
MD
Area de Microbiología, Facultad de Medicina, Universidad de Oviedo, Spain.
Méndez
F J
FJ
Vázquez
F
F
del Mar García-Suárez
M
M
de los Toyos
J R
JR
D001426:Bacterial Proteins; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D008163:Luminescent Measurements; D011994:Recombinant Proteins / Q000032:analysis; D012680:Sensitivity and Specificity; D013301:Streptolysins / Q000032:analysis* / Q000097:blood
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae