Sven Bocklandt, Ph.D. Species Director and Anthony Mastracci IV, Associate Researcher from Colossal Biosciences recently joined Cell Microsystems for a webinar titled “Reviving the past, safeguarding the future: challenges and innovations in cell culture.”
In many labs across the world, fluorescence-activated cell sorters (FACS) have remained the gold standard instrumentation for cell line development and gene editing workflows. While FACS is perhaps the most well-established automated methodology, cell sorters can present technical challenges such as:
In addition, for many difficult-to-culture or sensitive cell types, high-pressure fluidics-based sorting can lead to loss of cell viability, phenotypic drift, and the inability to recover rare cells.
Faced with some of these changes, Colossal Biosciences looked for an alternative technology that would augment their flow-based workflows and alleviate these pain points. Colossal discovered CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology, a gentle and cost-effective solution for their issues.
In this webinar, Colossal scientists describe the changes to their workflow upon integrating CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology and discuss new avenues that were not previously attainable with FACS. Take a look below as Dr. Sven Bocklandt and Anthony Mastracci IV explain how CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology automated and instilled confidence in their workflows.
To isolate cells to make monoclonal cell lines, the standard in the field really is to use FACS or fluorescence activated cell sorting. That puts several stresses on these cells. They sit in a sorting buffer sometimes for a long time, they're exposed to high pressure during the sorting, and then importantly they end up alone in a well without the ability to secrete and share growth factors with other cells.
We are using the CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More AIR, from Cell Microsystems to isolate cells in a way that is more gentle than FAC sorting. We seed cells onto a culture plate that is made up of thousands of microscopic rafts where they share the media and thus they can keep exchanging growth factors as if they are in a standard cell culture dish. We were concerned about cells transferring between wells, but we’ve been able to confirm the cells maintain monoclonalityEvidence that a colony/line originated from one cell; often supported by time-course imaging and traceability. More.
Cloning mouse embryonic stem cells is a tedious process of spending hours hunched over a stereoscope, meticulously using a P20 pipette to manually pick clones from a 6-well flask or larger. It is quite labor-intensive and places a significate strain on our scientists, consequently diverting their attention from higher-level tasks.
Upon implementing the CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More, we immediately noticed its ability to automatically pick clones of interest. This, in turn, freed up our scientists to focus on other lab tasks. Now the ability to observe individual ESLs just four hours after seeding expands our editing workflows and instils greater confidence in our research. This enables us to explore new avenues that were previously inaccessible.
Our approach involved seeding ESLs at low density on a six-well plate, followed by waiting seven to nine days before examining under the stereoscope. This method occasionally resulted in the formation of polyclonal lines when two ESLs either landed in close proximity to one another or failed to fully dissociate appearing as a single colony days later. Consequently, screening for polyclonal during genotyping became necessary.
Now using CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology, we have the ability to visualize single ES cells just four hours post-seeding possessing greater confidence in the monoclonal nature of colonies being generated. And as matter of fact, since implementing the CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More, we have yet to encounter a polyclonal ES colony.
mRNA has a short editing window peaking at around eight hours and tapering off over the course of 48 hours visualizing early ESL transfections. We can see the tapering off of the fluorescence begining to fade resulting in low SNR. This now demonstrates to us that we can start using fluorescence for positive selection for mouse ESLs.
Upon implementing CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology, Colossal automated their clone picking, noticed cells were more viable within their natural environment, and could visualize earlier within the short editing window. They were able to avoid excessive costly screening, having confirmed monoclonal colonies in their hands. These advantages demonstrate the flexibility, ease of use, and confidence Colossal has in the CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology. Colossal now has the ability to accelerate their research freeing up scientists from constantly sorting, re-analyzing, or additionally screening their cells.
To see more information on the technology that helped Colossal accelerate their workflows and enable their scientists to focus their attention back to higher-level tasks, visit our website and read about CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Technology.
Lexi Land contributes to developing scientific workflows that are compatible with the CellRaft® AIR System. Her focus at Cell Microsystems involves stem cell research, 3D organoid work, as well as adherent and suspension cell culture. Lexi received a Bachelor of Science in Biological Sciences from North Carolina State University with a focus in molecular, cellular, and developmental biology.
By Jessica Hartman, Ph.D.
Toiling in the lab
With a deadline on its way
To the scope I go
On a Saturday
I don’t see a clone
I’ve looked for one that’s right
What misery these data bring,
I’ll have to look all night,
Oh! Single cells, single cells, dilutions aren’t the way
Oh, I wish I had an AIR to pick a clone, to-da-ay!
Single cells, single cells, I need a better way
Use a Raft to grow a clone or be sad this holiday!
Use a Raft to grow a clone or be sad this holiday!
Now it’s getting late
My spirits are so low
My PI will hate
If my cells don’t grow
My eyesight’s getting dim
A clone I cannot see
My chances are so slim
I need CellRaftA microscale polystyrene growth surface within an array used to spatially segregate cells/colonies while maintaining shared media access, enabling imaging over time and targeted isolation. More Cytometry
Oh! Single cells, single cells, sorters aren’t the way
Oh, I wish I had an AIR to pick a clone, to-da-ay!
Single cells, single cells, I need a better way
Use a Raft to grow a clone or be sad this holiday!
Use a Raft to grow a clone or be sad this holiday!
Oh! Single cells, single cells, dispensers aren’t the way
Oh, I wish I had an AIR to pick a clone, to-da-ay!
Single cells, single cells, I need a better way
Use a Raft to grow a clone or be sad this holiday!
Use a Raft to grow a clone or be sad this holiday!
“Single Cell” vocalist: Virginia Laurie
