Microfluidics can improve epigenomic analysis, team says

Chang Lu, PhD, (left) and

his student, Zhenning Cao.

Dr Lu is holding the chip

used in the study.

Photo from Virginia Tech

A new technique enables epigenomic analysis using fewer cells than other methods involving chromatin immunoprecipitation and deep sequencing (ChIP-seq), according to researchers.

The technique, microfluidic oscillatory washing-based ChIP-seq (MOWChIP-seq), allows for genome-wide analysis of histone modifications using as few as 100

cells.

Using MOWChIP-seq, researchers uncovered new information regarding early hematopoiesis.

The team described this work in Nature Methods.

They used multilayer soft lithography to create a poly (dimethylsiloxane) device with a microfluidic chamber for ChIP. The researchers flowed magnetic beads coated with a ChIP antibody into the chamber, which formed a packed bed.

They then flowed sonicated chromatin fragments through the chamber, which were adsorbed onto the bead surface. The team said the gaps between the immunoprecipitation beads are smaller than 2 μm and facilitate rapid, high-efficiency adsorption of target chromatin fragments under the small diffusion length.

The researchers washed the beads by oscillatory washing in two different buffers to remove nonspecifically adsorbed chromatin fragments. Then, they flowed the beads out of the chamber and collected them for off-chip processing.

“The use of a packed bed of beads for ChIP allowed us to collect the chromatin fragments with a very high efficiency,” said study author Chang Lu, PhD, of Virginia Tech in Blacksburg.

“At the same time, effective washing for removing undesired molecules and debris guarantees the purity of the collected molecules. These two factors constitute a successful strategy for epigenomic analysis with extremely high sensitivity.”

In addition, the entire MOWChIP-seq process takes about 90 minutes.

To test MOWChIP-seq, Dr Lu and his colleagues used the technique to study the epigenomes of hematopoietic stem and progenitor cells (HSPCs) isolated from the fetal liver of a mouse.

“Little is known about the dynamics of the epigenome during embryonic hematopoiesis, largely due to the difficulty in isolating sufficient quantities of these cells from developing embryos,” said study author Kai Tan, PhD, of the University of Iowa in Iowa City. “This technology is the perfect tool for tackling this problem.”

MOWChIP-seq revealed new enhancers and super enhancers in the HSPCs.

By comparing all of the enhancers they identified to an enhancer catalog covering 16 blood cell types, the researchers found that 2561 (58%) of the enhancers they found were unique to fetal liver HSPCs. They said this suggests enhancer activity is highly dynamic during early hematopoiesis.

Now, the researchers plan to use MOWChIP-seq to study other epigenomic changes involved in inflammation and cancer.

“Our technology paves the way for studies of epigenomes with extremely low numbers of cells from animals and from patients,” Dr Lu said.

Virginia Tech Intellectual Properties has filed a utility patent on MOWChIP-seq on behalf of Dr Lu.

Chang Lu, PhD, (left) and

his student, Zhenning Cao.

Dr Lu is holding the chip

used in the study.

Photo from Virginia Tech

A new technique enables epigenomic analysis using fewer cells than other methods involving chromatin immunoprecipitation and deep sequencing (ChIP-seq), according to researchers.

The technique, microfluidic oscillatory washing-based ChIP-seq (MOWChIP-seq), allows for genome-wide analysis of histone modifications using as few as 100

cells.

Using MOWChIP-seq, researchers uncovered new information regarding early hematopoiesis.

The team described this work in Nature Methods.

They used multilayer soft lithography to create a poly (dimethylsiloxane) device with a microfluidic chamber for ChIP. The researchers flowed magnetic beads coated with a ChIP antibody into the chamber, which formed a packed bed.

They then flowed sonicated chromatin fragments through the chamber, which were adsorbed onto the bead surface. The team said the gaps between the immunoprecipitation beads are smaller than 2 μm and facilitate rapid, high-efficiency adsorption of target chromatin fragments under the small diffusion length.

The researchers washed the beads by oscillatory washing in two different buffers to remove nonspecifically adsorbed chromatin fragments. Then, they flowed the beads out of the chamber and collected them for off-chip processing.

“The use of a packed bed of beads for ChIP allowed us to collect the chromatin fragments with a very high efficiency,” said study author Chang Lu, PhD, of Virginia Tech in Blacksburg.

“At the same time, effective washing for removing undesired molecules and debris guarantees the purity of the collected molecules. These two factors constitute a successful strategy for epigenomic analysis with extremely high sensitivity.”

In addition, the entire MOWChIP-seq process takes about 90 minutes.

To test MOWChIP-seq, Dr Lu and his colleagues used the technique to study the epigenomes of hematopoietic stem and progenitor cells (HSPCs) isolated from the fetal liver of a mouse.

“Little is known about the dynamics of the epigenome during embryonic hematopoiesis, largely due to the difficulty in isolating sufficient quantities of these cells from developing embryos,” said study author Kai Tan, PhD, of the University of Iowa in Iowa City. “This technology is the perfect tool for tackling this problem.”

MOWChIP-seq revealed new enhancers and super enhancers in the HSPCs.

By comparing all of the enhancers they identified to an enhancer catalog covering 16 blood cell types, the researchers found that 2561 (58%) of the enhancers they found were unique to fetal liver HSPCs. They said this suggests enhancer activity is highly dynamic during early hematopoiesis.

Now, the researchers plan to use MOWChIP-seq to study other epigenomic changes involved in inflammation and cancer.

“Our technology paves the way for studies of epigenomes with extremely low numbers of cells from animals and from patients,” Dr Lu said.

Virginia Tech Intellectual Properties has filed a utility patent on MOWChIP-seq on behalf of Dr Lu.

Chang Lu, PhD, (left) and

his student, Zhenning Cao.

Dr Lu is holding the chip

used in the study.

Photo from Virginia Tech

A new technique enables epigenomic analysis using fewer cells than other methods involving chromatin immunoprecipitation and deep sequencing (ChIP-seq), according to researchers.

The technique, microfluidic oscillatory washing-based ChIP-seq (MOWChIP-seq), allows for genome-wide analysis of histone modifications using as few as 100

cells.

Using MOWChIP-seq, researchers uncovered new information regarding early hematopoiesis.

The team described this work in Nature Methods.

They used multilayer soft lithography to create a poly (dimethylsiloxane) device with a microfluidic chamber for ChIP. The researchers flowed magnetic beads coated with a ChIP antibody into the chamber, which formed a packed bed.

They then flowed sonicated chromatin fragments through the chamber, which were adsorbed onto the bead surface. The team said the gaps between the immunoprecipitation beads are smaller than 2 μm and facilitate rapid, high-efficiency adsorption of target chromatin fragments under the small diffusion length.

The researchers washed the beads by oscillatory washing in two different buffers to remove nonspecifically adsorbed chromatin fragments. Then, they flowed the beads out of the chamber and collected them for off-chip processing.

“The use of a packed bed of beads for ChIP allowed us to collect the chromatin fragments with a very high efficiency,” said study author Chang Lu, PhD, of Virginia Tech in Blacksburg.

“At the same time, effective washing for removing undesired molecules and debris guarantees the purity of the collected molecules. These two factors constitute a successful strategy for epigenomic analysis with extremely high sensitivity.”

In addition, the entire MOWChIP-seq process takes about 90 minutes.

To test MOWChIP-seq, Dr Lu and his colleagues used the technique to study the epigenomes of hematopoietic stem and progenitor cells (HSPCs) isolated from the fetal liver of a mouse.

“Little is known about the dynamics of the epigenome during embryonic hematopoiesis, largely due to the difficulty in isolating sufficient quantities of these cells from developing embryos,” said study author Kai Tan, PhD, of the University of Iowa in Iowa City. “This technology is the perfect tool for tackling this problem.”

MOWChIP-seq revealed new enhancers and super enhancers in the HSPCs.

By comparing all of the enhancers they identified to an enhancer catalog covering 16 blood cell types, the researchers found that 2561 (58%) of the enhancers they found were unique to fetal liver HSPCs. They said this suggests enhancer activity is highly dynamic during early hematopoiesis.

Now, the researchers plan to use MOWChIP-seq to study other epigenomic changes involved in inflammation and cancer.

“Our technology paves the way for studies of epigenomes with extremely low numbers of cells from animals and from patients,” Dr Lu said.

Virginia Tech Intellectual Properties has filed a utility patent on MOWChIP-seq on behalf of Dr Lu.