As you know, the M. hyopneumoniae ELISA kit from Oxoïd/Dako will no longer be available for several months. Among the options we propose to replace this test, there is the M. hyopneumoniae MFIA, as well as the PRRSV type 1 – PRRSV type 2 – M. hyopneumoniae MFIA for those who want to test for PRRSV antibodies in addition to M. hyopneumoniae antibodies.
MFIA is the acronym for Multiplexed Fluorescing Immuno Assay. MFIAs are immunoassays that use different kinds of microbeads (diameter: 5 μm), each with a specific fluorescent label.
Microbeads of a given kind are coated with a particular antigen. For example, in the case of the PRRSV type 1 – PRRSV type 2 – M. hyopneumoniae MFIA, three types of beads are used:
- beads A coated with PRRSV type 1 antigen
- beads B coated with PRRSV type 2 antigen
- beads C coated with M. hyopneumoniae antigen
MFIA tests are conducted as follows:
- Step 1: the sera to be tested are exposed to the mixture of the different beads in the wells of a microplate. During incubation (60 minutes at room temperature), the antibodies eventually present bind to the corresponding antigens.
- Step 2: the beads are then exposed to biotinylated anti-porcine IgG antibodies. During incubation (30 minutes at room temperature), these antibodies bind to the IgG antibodies which are bound to the corresponding antigens.
- Step 3: the beads are finally exposed to phycoerythrin (a fluorochrome) conjugated to streptavidin. During incubation (30 minutes at room temperature), streptavidin binds to the biotin of the biotinylated anti-porcine IgG antibodies.
- Step 4: at the end of step 3, the beads are examined with a fluorescence plate reader able of differentiating the different kinds of beads and, for each of them (each antigen), determining the intensity of the fluorescence generated by phycoerythrin.
Between steps, the beads are washed several times to eliminate all the elements that did not attach to them.
- Step 5: all the data are finally processed by a software which, for each tested serum and each antigen included in the test, compares the intensity of the fluorescence with that of positive and negative control sera to generate an index S/P (i.e. corrected sample fluorescence intensity / corrected positive control fluorescence intensity). If the S/P value is greater than or equal to a threshold value, the serum is considered positive for the corresponding antibodies; if it is lower, it is considered negative.
See examples of results for the MFIA tests and the relative sensitivity and specificity assessment of the different tests:
You can download the request forms in interactive pdf format including these MFIA tests:
For further information, feel free to contact the technical support team.