Steven Santana was recently featured in the National Action Council for Minorities in Engineering (NACME), Inc. 2012 Annual Report. This report, entitled “American Engineers: The Key to Global Competitiveness,” can be found at http://nacme.org/user/docs/AR2012Final.pdf.
Recently, we published a review on theory and experiment for electrokinetics with soft and charged interfaces. Our paper discusses the governing physics, modeling, and experimental verifications of theories for electrokinetics on soft and charged interfaces. To codifiy this understanding, we connect the cognate electrokinetic phenomena using the electrokinetic coupling matrix.
A soft and charged interface between a rigid wall and pure fluid phase. Effects of the soft and charged interface on fluid velocity profiles for pressure (solid line) and electric field (dashed line) are shown at left, and effects on the local potential distribution are shown at right. (Enlarge by clicking on the image.)
Descriptions of soft and charge interfaces deviate from classical theories of electrokinetics by postulating a porous charge layer at the interface between the completely impermeable wall and pure fluid. In this interfacial region, a large amount of charge may be attached to the porous layer. Depending upon the permeability of the porous layer, immense amplifications of electroosmosis, streaming potential, and/or conductance may be realized. Changes in these phenomena are not explained using rigid-surface electrokinetic theories.
Descriptions of soft and charged layers are essential to understand and predict the behavior of systems with non-rigid interfaces bearing fixed charge. Soft and charged interfaces are present in a variety of systems, both synthetic (e.g., polymer exchange membrane, grafted polymer layers) and natural (e.g., cartilage, lipid membranes). We hope the theories and experiments discussed in the review provide researchers with a framework to interpret and plan results and experiments.
We recently published a review in a “Microfluidics and Minaturization” special issue of Electrophoresis. This work was a collaboration between myself, Jason, Alex, Steven, and Brian. Brian and Steven contributed knowledge on rare cell adhesion, Alex discussed fluid mechanics at the microscale, and Jason and I reviewed device designed strategies.
An obstacle array is used to induce cross-streamline motion, bringing target cells into contact with an immunocapture surface, while smaller non-target cells have relatively few collisions (represented by stars).
The article reviews biorheology, rare cell surface markers and adhesion models, as well as general transport phenomena at the microscale. Several design strategies, including micromixers, porous filtration systems, and obstacle arrays, are presented in a transport context. A key conclusion is that advection (i.e., motion of the fluid itself) dominates diffusion in most rare cell capture devices; a successful device is designed not by maximizing the ratio of surface area to volume, but by inducing cross-streamline motion to bring cells into contact with a capture surface.
This review summarizes the knowledge we have gained developing the GEDI microdevice, and we hope that other researchers will find it useful in the development of their own rare cell capture devices.
x-posted to Erica Pratt’s Blog
BCa: Breast Cancer
CTC: Circulating tumor cell. Read about what they are and why they’re important here.
Epithelial cell surface marker: A protein or receptor sticking out of the cell membrane of epithelial cells.
EMT: Epithelial-to-mesenchymal transition, where cells lose biomarkers associated with their organ of origin and become more stem cell-like.
SNR: Signal-to-noise ratio.
Tumor genetic heterogeneity has emerged as an effective biomarker of malignant processes1-4. However, limited access to tissue in solid tumors makes repeated sampling and tracking of tumor mutations infeasible. CTCs can serve as a “liquid biopsy”, allowing researchers to study genetic progression in real time. The paper I’m reviewing today, “Single Cell Profiling of Circulating Tumor Cells: Transcriptional Heterogeneity and Diversity from Breast Cancer Cell Lines” by Powell et al., demonstrates the utility of single-CTC genetic profiling5. The article is Open Access and available on PLoS ONE.
The technology used in this study is called the MagSweeper, developed by the Jeffrey Lab at Stanford. Magnetic beads were coated with an antibody targeting epithelial cell surface markers. These antibody-coated (i.e. immunomagnetic) beads were mixed into blood samples, resulting in cancer cells covered in beads, as shown in figure D. The blood samples were diluted with saline solution, and cancer cells were extracted using a magnetic source. Captured cells were washed while attached to the magnet, and then released when the applied field was removed, as shown in figure B. Cell gene expression and viability were shown to be unaffected by this capture process.
x-posted at Erica Pratt’s blog
Cell Surface Marker: Some protein or receptor sticking out of the cell membrane. I explain this more in depth in my immunocapture post.
CTC: Circulating tumor cell. Read about what they are and why they’re important here.
Microemboli: A small mass of cells or tissue inside the bloodstream.
Platelets: aka thrombocytes, important in the formation of blood clots.
Phenotype: Observable characteristics of a cell.
RBCs: Red blood cells, aka erythrocytes.
Senescence: When a cell hits the Hayflick limit and can no longer divide naturally.
WBCs: White blood cells, aka leukocytes.
In my “How to Sort CTCs” series, I covered a variety of sorting methodologies used for patient prognosis. However, before clinical implementation, it is important characterize device performance with a series of standards. This is impossible to do with a patient blood sample, because there is an unknown number of CTCs floating around with other blood cells, which can be effected by the cancer treatment process (e.g. radiation patients often have anemia)1. Furthermore, this is all changing dynamically as a function of both time and treatment.
For this reason, engineers need an alternative system that can serve as a patient blood model, but it is repeatable and controllable. Immortalized cancer cell lines—derived from cancers of various organs—are commonly used for this purpose. A normal human cell can only divide a set number of times before it undergoes senescence; this is called the Hayflick limit. Immortalized cell lines have been genetically altered to surpass the Hayflick limit and continue dividing indefinitely.
This enables researchers to create standardized systems to benchmark their technology with. A known number of cancer cells can be spiked in varying ratios with different blood components, allowing for measurement of sensitivity and specificity of capture, along with other metrics. For this reason…
x-posted at Erica Pratt’s Blog
Anion:a negatively charged ion.asophil, Lymphocyte, Neutrophil: Different types of white blood cells.
Biofouling: Fouling of surface with biological material.
Cation: A positively charged ion.
Conductivity: Measure of a substance’s ability to conduct electricity.
Cytoplasm: Inner contents of the cell, which holds everything outside the nucleus.
CTC: Circulating tumor cell. Read about what they are and why they’re important here.
Erythrocyte: Red blood cell.
PBMCs: Peripheral mononuclear blood cells, aka blood cells that have a nucleus (e.g. white blood cells). Cartoon of the different kinds of white blood cells here.
MDA-MB-***: Human breast carcinoma immortalized cell lines.
Phenotype: Observable characteristics of a cell.
This is the last major sorting technique in this series (for now), and I will be using the review paper I co-wrote with Charlie Huang as the framework for my description of this technique1. Charlie works extensively on electrokinetic manipulation of cells, and his half of the review paper lends itself well to explaining how electrokinetics can be used to sort CTCs.
Most CTC sorting devices target some observed cancer cell phenotype that was determined from studying tumor tissue directly, or from using immortalized cancer cell lines. This means that active sorting techniques, like size-based selection and immunocapture, require some level of a priori knowledge about CTCs before you can engineer a device to capture them. Microscopic characterization is one CTC identification method that circumvents this problem, fixing (killing) the cells, and then using imaging in combination with rapid scanning to look at almost everything present in the blood sample. Electrokinetic separation of cancer cells is another, but enables live cell isolation without knowing its physical or biochemical properties beforehand.
There are two commonly used types of electrokinetic manipulation for mammalian cells, electrophoresis (EP) and dielectrophoresis (DEP). Electrophoresis involves applying a uniform electric field across a charged particle, causing it to polarize (i.e. free charge aligns with the electric field), inducing a net particle migration. However, if a uniform electric field is applied to an electrically neutral particle, the charges will polarize to form a dipole, but there is no actuation because the force on each anion is cancelled out by the force on its respective cation, and vice versa. To induce actuation, a non-uniform electric field must be applied (Dielectrophoresis), causing the charge on one side of the particle to feel the electrical force more strongly than the other, resulting in particle migration—as shown in the example below (thanks to the Kirby Lab DEP subgroup for the great schematic!).
Electrophoresis is good for moving charged particles around; however, the net charge from cell type-to-cell type is often not distinct enough to sort cells with high resolution. In contrast, dielectrophoresis is excellent for sorting cells because motion is dependent not on net charge, but on cell membrane and cytoplasm electrical properties as well as cell size, as dictated by this equation:
I attended the recent Society of Hispanic Professional Engineers Annual Conference in Forth Worth, Texas, where I presented a talk entitled “Capture and Interrogation of Circulating Tumor Cells in Tissue-Engineered GEDI Microfluidic Devices.” This talk addressed a handful of gaps in current knowledge regarding cancer progression and the expectation that tissue-engineered GEDI technologies can help to address these issues.
I earned the Paper-Presentation Competition award for this work.
