MAIGRET: a CRISPR-based immunoassay that employs antibody-induced cell-free transcription of CRISPR guide RNA strands

Here we report on the development of a CRISPR-based assay for the sensitive and specific detection of antibodies and antigens directly in complex sample matrices. The assay, called MAIGRET (Molecular Assay based on antibody-Induced Guide-RNA Enzymatic Transcription), is based on the use of a responsive synthetic DNA template that trigger the cell-free in vitro transcription of a guide RNA strand upon recognition of a specific target antibody. Such transcribed guide RNA activates the DNA collateral activity of the Cas12 enzyme, leading to the downstream cleavage of a fluorophore/quencher-labelled reporter and thus resulting in an increase in the measured fluorescence signal. We have used MAIGRET for the detection of six different antibodies with high sensitivity (detection limit in the picomolar range) and specificity (no signal in the presence of non-target antibodies). MAIGRET can also be adapted to a competitive approach for the detection of specific antigens. With MAIGRET we significantly expand the scope and applicability of CRISPR-based sensing approaches to potentially enable the measurement of any molecular target for which an antibody is available.