Protocols

The experimental workflow for NTT-seq is very similar to running CUT&Tag, and we recommend using the excellent protocol provided by the CUT&Tag authors when running NTT-seq: https://doi.org/10.17504/protocols.io.bcuhiwt6

The following modifications are needed in comparison to the standard CUT&Tag workflow:

1. Stain cells or nuclei using a mixture of different primary antibodies for the different targets you’re interested in. Ensure that each primary antibody is able to be bound by a corresponding nanobody-Tn5 fusion protein.
2. Bind a pooled mixture of nb-Tn5 proteins.
3. For single-cell applications, wash cells or nuclei after tagmentation and proceed with the 10x Genomics scATAC-seq protocol steps following tagmentation.