An enzyme immunoassay for the qualitative and semiquantitative determination of IgM-class antibodies to Mycoplasma pneumoniae in serum.
The mycoplasms belong to the class Mollicutes comprising three distinct families and four genera, one of which is Mycoplasma with over 60 species. Mycoplasms are the smallest free living organisms known (300 to 500 nm in diameter) and unlike regular bacteria they lack a cell wall. Mycoplasms are extracellular parasites, especially on mucous membranes, which can cause infections in human, animals, plants, and cell cultures. Mycoplasma pneumoniae is primarily a respiratory pathogen (obligat) in human involving the nasopharynx, throat, trachea, bronchi, bronchioles, and alveoli. Other Mycoplasms, M. buccale, M. faucium, M. orale and M. salivarium are commensals in the oral cavity. Mycoplasma hominis and Ureaplasma urealyticum inhabit primarily the genital tract and may act as opportunistic invaders. M. pneumoniae is by far the most important pathogen of this group. Infection with M. pneumoniae occurs worldwide. Its epidemiology has been studied primarily in the USA, Europe, and Japan. Infections are endemic in larger urban areas, and epidemic increases are observed at varying intervals. M. pneumoniae has been estimated to cause 15-20% of all pneumoniae; the rate is highest in children and young adults. 74% of
infections with M. pneumoniae are asymptomatic, reinfection may occur. Naturally acquired immunity to infection with M. pneumoniae appears to be of limited duration (2-3 years).
The DRG Mycoplasma pneumoniae IgM ELISA Kit is a solid phase enzyme-linked immunosorbent assay (ELISA)Patient samples are diluted with Sample Diluent and additionally incubated with IgG-RF-Sorbent, containing hyper-immune anti-human IgG-class antibody to eliminate competitive inhibition from specific IgG and to remove rheumatoid factors. This pretreatment avoids false negative or false positive results. Microtiter wells as a solid phase are coated with Mycoplasma pneumoniae antigen. Pretreated patient specimens and ready-for-use controls are pipetted into these wells. During incubation Mycoplasma pneumoniae-specific antibodies of positive specimens and controls are bound to the immobilized antigens. After a washing step to remove unbound sample and control material horseradish peroxidase conjugated anti-human IgM antibodies are dispensed into the wells. During a second incubation this anti_IgM conjugate binds specifically to IgM antibodies resulting in the formation of enzyme-linked immune complexes. After a second washing step to remove unbound conjugate the immune complexes formed (in case of positive results) are detected by incubation with TMB substrate and development of a blue color. The blue color turns into yellow by stopping the enzymatic indicator reaction with sulfuric acid. The intensity of this color is directly proportional to the amount of Mycoplasma pneumoniae-specific IgM antibody in the patient specimen. Absorbance at 450 nm is read using an ELISA microtiter plate reader.