3️⃣ So what links this pre-activation repositioning to multipotency? We find it's closely tied to increased chromatin accessibility at sites bound by key limb transcription factors. This includes several factors shown to act together via a “coordinator motif” doi.org/10.1016/j.ce...

2️⃣ But not only genes reposition. Genes escape the periphery together with the surrounding enhancers and TADs that control their expression. Using polymer modeling & mutants, we show that CTCF boundaries and 3D chromatin topology prevent this repositioning from spreading to neighboring regions.

1️⃣ At the start of limb formation, hundreds of genes move away from the repressive nuclear periphery in multipotent progenitors. Amazingly, this repositioning can precede gene activation by up to four developmental days (E9.5 -> E13.5), suggesting it is a key priming step for multipotency!

Developmental genes have really precise expression in time and space and so must spend most of their time inactive. So how and when do they "wake up" to perform their function? Leveraging scDam&T-seq in embryonic limb development, we uncover three exciting new insights:

Is your rim staining real, or Ab-trapping? Solutions:
✅ Increase incubation time
✅ Tissue sections (antigen retrieval)
✅ Use lower-affinity antibodies/nanobodies
✅ Check cell-to-cell expression: rims only in high-expression cells = likely artifact

This artifact can impact any assay that relies on antibody diffusion, such as classic fluorescence microscopy, but also new-generation genomics techniques, such as CUT&Tag and CUT&RUN - SO WATCH OUT. See falsely peripheral H3K9Me2 Cut&Tag in mouse rod cells 👇

Ab-trapping occurs when antibodies fail to penetrate due to:
1️⃣ High epitope abundance
2️⃣ High antibody affinity
3️⃣ Low diffusion rate
Any one factor is enough! Potentially affects many antibody–epitope pairs under the right conditions. Examples 👇

Is your rim staining real, or Ab-trapping? Solutions:
✅ Increase incubation time
✅ Tissue sections (antigen retrieval)
✅ Use lower-affinity antibodies/nanobodies
✅ Check cell-to-cell expression: rims only in high-expression cells = likely artifact

This artifact can impact any assay that relies on antibody diffusion, such as classic fluorescence microscopy, but also new-generation genomics techniques, such as CUT&Tag and CUT&RUN - SO WATCH OUT. See falsely peripheral H3K9Me2 Cut&Tag in mouse rod cells 👇

Ab-trapping occurs when antibodies fail to penetrate due to:
1️⃣ High epitope abundance
2️⃣ High antibody affinity
3️⃣ Low diffusion rate
Any one factor is enough! Potentially affects many antibody–epitope pairs under the right conditions. Examples 👇