Development of a Microfluidic DEP-enhanced DLD Method for Sorting Bone Marrow Adipocytes

Abstract

Isolating fragile and heterogeneous cell populations from complex tissues remains a core challenge in biomedical research. In the bone marrow (BM), bone marrow adipocytes (BMAds) are key regulators of hematopoiesis and metabolism, yet their large size, fragility, and lack of specific surface markers have hindered their isolation using conventional methods such as fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS), which often compromise cell viability or lack specificity for lipid-rich cells. Here, we introduce a label-free microfluidic platform that integrates deterministic lateral displacement (DLD) with dielectrophoresis (DEP) to enable high-precision, tunable sorting of BM-derived cells based on size and dielectric properties. In the first phase, we established a clog-free DLD workflow by optimizing sample preparation of murine and human BM aspirates through low-speed centrifugation and lipid layer removal. Although mature BMAds were underrepresented, the platform enabled continuous processing of heterogeneous BM populations without clogging or damage. To overcome the fixed critical size inherent to standard DLD, we incorporated planar electrodes into the micropost array, creating a hybrid DEP-DLD device with voltage-controlled sorting thresholds comparable to an electrically adjustable sieve. Increasing the voltage allows smaller particles or cells to be deflected and sorted. By applying different voltage regions across the chip, multifraction separation of 10, 20, and 30 μm polystyrene beads was achieved with >90% sorting purity across three outlets. Biological validation using OP9 stromal cells and differentiated adipocytes confirmed cellular responsiveness to negative DEP forces at 100 kHz and 6.5 Vpp. However, while cells were repelled from the electrodes, the applied DEP force was insufficient to fully displace them into lateral trajectories. In the future, this DEP-DLD platform offers a non-destructive, tunable alternative to conventional sorting technologies, enabling the isolation of fragile BM-derived cells and the recovery of pure populations for downstream functional analysis in health and disease.

Popular Abstract

Sorting the unsortable: A new way to gently sort bone marrow cells

Hidden deep inside our bones is a surprisingly lively world, a miniature city where blood-producing cells, immune cells and fat cells share space, signals and secrets. Among these residents, bone marrow adipocytes (BMAds) stand out as the mysterious introverts: large, fragile fat cells that scientists believe may influence everything from blood development to bone strength and metabolic disease. But while they may hold big answers, they are almost impossibly delicate. Try to isolate one, and it bursts like a soap bubble. Traditional cell-sorting methods are not much kinder. Many rely on surface markers, molecular “ID tags”, on the cell membrane that BMAds simply sparsely express, and the mechanical stress involved is catastrophic for a cell with the structural resilience of wet tissue paper. Most adipocytes never make it out alive.

The challenge was simple to ask but tricky to solve: could we sort these cells without squeezing, staining, or destroying them? This is where microfluidics enters the story, a technology that guides cells through hair-thin channels using smooth flows and gentle forces. The platform developed in this project uses two such mechanisms working in tandem. The first, Deterministic Lateral Displacement (DLD), functions like a microscopic obstacle course where small cells slip straight through while larger cells drift off to the side. The second, Dielectrophoresis (DEP), separates cells using subtle electrical forces, allowing us to tune how they move simply by turning a voltage dial.

The ultimate goal is to isolate BMAds reliably so we can finally study them properly to learn whether they are helpful supporters of blood formation, silent disruptors of bone health, or something far more complex. And the early steps are promising. The DLD system could process real bone marrow samples without clogging or destroying cells, and when DEP was added to the mix, we could sort synthetic particles into size-defined groups with impressive accuracy just by adjusting the electrical settings. Most importantly, BMAds survived the experience but did not respond as anticipated. With further optimisation and more refined tuning, this technology may finally give us the ability to capture these elusive cells intact.

If that happens, a door opens. We could start asking the questions that have long been out of reach: How do BMAds regulate blood production? What is their role in cell metabolism? How are they linked to blood and bone diseases? Unlocking these answers might reshape our understanding of bone marrow biology and perhaps change the way we sort clinical samples in the future.

• author: Peltonen, Marina
• supervisor: Maria Nordin
• organization: Division for Biomedical Engineering
• alternative title: Utveckling av en mikrofluidisk DEP-förstärkt DLD-metod för separation av benmärgsadipocyter
• course: BMEM01 20252
• year: 2026
• type: H2 - Master's Degree (Two Years)
• subject: Technology and Engineering
• keywords: Microfluidics, Deterministic lateral displacement, Dielectrophoresis, Label-free sorting, Bone marrow, Adipocytes, Cell sorting, Microengineering, Microfabrication, Lab-on-a-chip
• language: English
• additional info: 2025-23
• id: 9218223
• date added to LUP: 2026-02-03 12:41:30
• date last changed: 2026-02-03 12:41:30