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As previously discussed, multiplexing is assaying several different genes in the same reaction. Multiplexing has become possible due the high specificity of probe design. Development of multiplexing PCR reactions can be complicated because it has to be ensured that the probes do not interfere or react with each other. The process can also be time-consuming because each reaction may require different conditions. Figuring out the ideal mix for all the probes can take time. When developing a new multiplex test, it is important to weigh the time involved and costs against running each PCR reaction individually.

There are many pre-established multiplex tests that can be purchased and used in laboratories today. One example is a multiplex PCR test that can distinguish between Staphylococcus aureus and coagulase-negative Staphylococcus and determine the presence of the gene that is resistant to oxacillin. Another example is a multiplex PCR test that can detect all four species of Plasmodium, the cause of malaria.