Your CUT&RUN Library Has Peaks.
But How Do You Know They Are Real?

Successful library generation is only part of a chromatin profiling experiment. Reliable interpretation also requires a way to distinguish target-specific enrichment from assay background. That makes IgG negative controls and carefully selected secondary antibodies important components of CUT&RUN and CUT&Tag workflows.

CUT&RUN and CUT&Tag Profile Chromatin in Different Ways

Both methods are used to map histone modifications and chromatin-associated proteins across the genome.

In CUT&RUN, an antibody directs a protein A/G-MNase fusion toward the target, allowing DNA near antibody-bound chromatin to be cleaved and released.

CUT&Tag instead recruits a protein A-Tn5 or related transposase complex, which performs targeted tagmentation around the antibody-bound site.

The Challenge: Background

A sequencing peak does not automatically represent true biological enrichment. Non-specific antibody interactions, enzyme recruitment and other sources of background can generate reads that appear in the final library.

A properly designed negative control provides the reference needed to distinguish target-specific enrichment from assay background.

The Role of an IgG Negative Control in CUT&RUN

A non-specific IgG matched to the experimental antibody host can be processed alongside experimental samples.

Because the control should not recognize the biological target, its sequencing profile provides an estimate of non-specific background generated by the workflow.

Clean separation between experimental enrichment and IgG background supports more confident downstream peak calling.

In CUT&Tag, the Secondary Antibody Matters Too

For rabbit primary antibodies, a compatible secondary antibody can increase the number of protein A interaction sites associated with the bound primary, supporting recruitment of the Tn5 complex.

However, non-specific secondary antibody interactions can also recruit the tagmentation machinery away from the intended target.

Secondary antibody specificity therefore contributes directly to the signal-to-background performance of the workflow.

Guinea Pig anti-Rabbit IgG — ABIN101961

ABIN101961 is a preadsorbed polyclonal Guinea Pig anti-Rabbit IgG (Heavy & Light Chain) antibody.

The same reagent has been independently evaluated for two distinct roles in chromatin profiling workflows.

Why Preadsorption Matters

Cross-reactivity with immunoglobulins or serum proteins from unintended species can increase background in antibody-based workflows.

ABIN101961 is solid-phase preadsorbed, and manufacturer testing reports no observed reaction with human, mouse or goat serum proteins.

The product is listed for both CUT&RUN and CUT&Tag, with a general recommended dilution of 1:100 for each application.

CUT&RUN / CUT&Tag Checklist

✓ Is the primary antibody validated for the intended application?

✓ Is an appropriate IgG negative control included?

✓ Is a positive histone control included?

✓ Is the primary antibody host compatible with the secondary?

✓ Has secondary antibody cross-reactivity been considered?

✓ Does the negative-control library show sufficiently low background?

✓ Is the control dataset incorporated appropriately during peak calling?

Planning a CUT&RUN or CUT&Tag Experiment?

Dana Bioscience can help source primary antibodies, IgG controls, secondary antibodies, magnetic beads and other reagents for CUT&RUN and CUT&Tag workflows.

Share your target or histone mark, sample species, primary antibody host and workflow, and we can help identify appropriate research reagents.