Although ELISA and flow cytometry have been the traditional assays of choice for antibody screening, they have many disadvantages particularly for cell surface antigens expressed at low levels. To address these limitations, homogeneous or “mix-and-read” assays have been developed and coupled with appropriate high sensitivity detection systems.
The high sensitivity of this approach allows screening of supernatants much earlier than ELISA or flow cytometry, using as little as 5 μL of sample. The method also reduces reagent requirements which is beneficial in the study of high cost or low abundance antigens.
mirrorball (TTP Labtech) and the ABI 8200 (Applied Biosystems) – also known as FMAT – contain specific design features for the analysis of homogeneous binding assays. They offer high sensitivity through the use of laser excitation and PMT detection, which allows the detection of low affinity antibodies and low abundance antigens. Both systems have optics that exclude background fluorescence during scanning to dramatically increase the signal to noise ratio on the relevant target.
This study comparing mirrorball and the ABI 8200 demonstrates that the detection sensitivity of mirrorball is comparable to that of the ABI 8200. Furthermore transfer of existing ABI 8200 protocols proved straightforward and required minimal development to run on a mirrorball.