---
title: "Why Is There High Background in My CUT&RUN Sequencing Data?"
slug: "why-is-there-high-background-in-my-sequencing-data"
description: "Troubleshoot high background in CUT&RUN sequencing data by evaluating sample prep, antibodies, MNase digestion, library prep, and QC."
updated: 2024-10-01T17:28:04Z
published: 2024-10-01T17:28:04Z
canonical: "support.epicypher.com/why-is-there-high-background-in-my-sequencing-data"
---

> ## Documentation Index
> Fetch the complete documentation index at: https://support.epicypher.com/llms.txt
> Use this file to discover all available pages before exploring further.

# Why is there high background in my sequencing data?

High background in sequencing data could indicate poor sample or library prep, over-digestion by MNase, DNA damage, or antibody failure.

To troubleshoot this, use 500,000 cells per reaction, including reactions with control antibodies and the K-MetStat Panel. Read the tips below:

- [Confirm sample prep](/v1/docs/confirm-high-quality-sample-prep) and include all [recommended controls and success metrics](/v1/docs/summary-of-cut-and-run-quality-control-and-success-metrics).
- Process cells quickly and resuspend in cold Antibody Buffer.
- Test multiple antibodies to experimental target.
- Ensure MNase digestion is incubated at 4˚C for 2 hours.
- Keep adapter ligation reagents on ice during ligation setup.
- Double check that library prep PCR parameters are accurate.