Rebecca L. Matz

Cell volume changes during apoptosis monitored in real time using digital holographic microscopy

Cellular volume changes play important roles in many processes associated with the normal cell activity, as well as various diseases. Consequently, there is a considerable need to accurately measure volumes of both individual cells and cell populations as a function of time. In this study, we have monitored cell volume changes in real time during apoptosis using digital holographic microscopy. Cell volume changes were deduced from the measured phase change of light transmitted through cells. Our digital holographic experiments showed that after exposure to 1 μM staurosporine for 4 h, the volumes of KB cells were reduced by ~50-60%, which is consistent with previous results obtained using electronic cell sizing and atomic force microscopy. In comparison with other techniques, digital holographic microscopy is advantageous because it employs noninvasive detection, has high time resolution, real time measurement capability, and the ability to simultaneously investigate time-dependent volume changes of both individual cells and cell populations.

Challenge faculty to transform STEM learning

Focus on core ideas, crosscutting concepts, and scientific practices Models for higher education in science, technology, engineering, and mathematics (STEM) are under pressure around the world. Although most STEM faculty and practicing scientists have learned successfully in a traditional format, they are the exception, not the norm, in their success. Education should support a diverse population of students in a world where using knowledge, not merely memorizing it, is becoming ever more important. In the United States, which by many measures is a world leader in higher education, the Presidents Council of Advisors on Science and Technology (PCAST) recommended sweeping changes to the first 2 years of college, which are critical for recruitment and retention of STEM students (1). Although reform efforts call for evidence-based pedagogical approaches, supportive learning environments, and changes to faculty teaching culture and reward systems, one important aspect needs more attention: changing expectations about what students should learn, particularly in college-level introductory STEM courses. This demands that faculty seriously discuss, within and across disciplines, how they approach their curricula.

Dual-wavelength linear regression phase unwrapping in three-dimensional microscopic images of cancer cells

We present a study of the three-dimensional structure of cancer cells using dual-wavelength phase-imaging digital holographic microscopy. Phase imaging of objects with optical height variation greater than the wavelength of light is ambiguous and causes phase wrapping. By comparing two phase images recorded at different wavelengths, the images can be accurately unwrapped. The unwrapping method is computationally fast and straightforward, and it can process complex topologies. Additionally, the limitations on the total optical height are significantly relaxed. This new methodology is widely applicable to other phase-imaging techniques as well as in applications beyond optical microscopy.

Characterizing college science assessments: The three-dimensional learning assessment protocol

Many calls to improve science education in college and university settings have focused on improving instructor pedagogy. Meanwhile, science education at the K-12 level is undergoing significant changes as a result of the emphasis on scientific and engineering practices, crosscutting concepts, and disciplinary core ideas. This framework of “three-dimensional learning” is based on the literature about how people learn science and how we can help students put their knowledge to use. Recently, similar changes are underway in higher education by incorporating three-dimensional learning into college science courses. As these transformations move forward, it will become important to assess three-dimensional learning both to align assessments with the learning environment, and to assess the extent of the transformations. In this paper we introduce the Three-Dimensional Learning Assessment Protocol (3D-LAP), which is designed to characterize and support the development of assessment tasks in biology, chemistry, and physics that align with transformation efforts. We describe the development process used by our interdisciplinary team, discuss the validity and reliability of the protocol, and provide evidence that the protocol can distinguish between assessments that have the potential to elicit evidence of three-dimensional learning and those that do not.

Polyplex Exposure Inhibits Cell Cycle, Increases Inflammatory Response, and Can Cause Protein Expression Without Cell Division

We sought to evaluate the relationship between cell division and protein expression when using commercial poly(ethylenimine) (PEI)-based polyplexes. The membrane dye PKH26 was used to assess cell division, and cyan fluorescent protein (CFP) was used to monitor protein expression. When analyzed at the whole population level, a greater number of cells divided than expressed protein, regardless of the level of protein expression observed, giving apparent consistency with the hypothesis that protein expression requires cells to pass through mitosis in order for the transgene to overcome the nuclear membrane. However, when the polyplex-exposed population was evaluated for the amount of division in the protein-expressing subpopulation, it was observed that substantial amounts of expression had occurred in the absence of division. Indeed, in HeLa S3 cells, this represented the majority of expressing cells. Of interest, the doubling time for both cell lines was slowed by ~2-fold upon exposure to polyplexes. This change was not altered by the origin of the plasmid DNA (pDNA) transgene promoter (cytomegalovirus (CMV) or elongation factor-1 alpha (EF1α)). Gene expression arrays in polyplex-exposed HeLa S3 cells showed upregulation of cell cycle arrest genes and downregulation of genes related to mitosis. Chemokine, interleukin, and toll-like receptor genes were also upregulated, suggesting activation of proinflammatory pathways. In summary, we find evidence that a cell division-independent expression pathway exists, and that polyplex exposure slows cell division and increases inflammatory response.

Dual-wavelength digital holographic imaging with phase background subtraction

Three-dimensional digital holographic microscopic phase imaging of objects that are thicker than the wavelength of the imaging light is ambiguous and results in phase wrapping. In recent years, several unwrapping methods that employed two or more wavelengths were introduced. These methods compare the phase information obtained from each of the wavelengths and extend the range of unambiguous height measurements. A straightforward dual-wavelength phase imaging method is presented which allows for a flexible tradeoff between the maximum height of the sample and the amount of noise the method can tolerate. For highly accurate phase measurements, phase unwrapping of objects with heights higher than the beat (synthetic) wavelength (i.e. the product of the original two wavelengths divided by their difference), can be achieved. Consequently, three-dimensional measurements of a wide variety of biological systems and microstructures become technically feasible. Additionally, an effective method of removing phase background curvature based on slowly varying polynomial fitting is proposed. This method allows accurate volume measurements of several small objects with the same image frame.

Dual wavelength digital holography phase unwrapping by linear regression

Digital holography records the superposition of the object and reference waves. The subsequent reconstruction of both amplitude and phase of the optical field is done by numerically propagating the optical field along the direction perpendicular to the hologram plane in accordance with the laws of diffraction. Phase changes undergone by a light wave passing through or reflecting from objects can be converted to the optical thickness or height measurements, providing the three dimensional structural information about the object. Our dual wavelength phase imaging method allows three dimensional measurements of a wide variety of biological systems and microstructures.

Digital Holographic Microscopy Study of Early Morphological Changes during Apoptosis

We present a digital holographic study of cellular volume changes during apoptosis. The reconstruction is performed by the angular spectrum method. The phase unwrapping is done in software using our varying reconstruction distance algorithm.

Evaluating the extent of a large-scale transformation in gateway science courses

An institutional effort to transform gateway science courses is evaluated using a novel approach based on course assessments. We evaluate the impact of an institutional effort to transform undergraduate science courses using an approach based on course assessments. The approach is guided by A Framework for K-12 Science Education and focuses on scientific and engineering practices, crosscutting concepts, and core ideas, together called three-dimensional learning. To evaluate the extent of change, we applied the Three-dimensional Learning Assessment Protocol to 4 years of chemistry, physics, and biology course exams. Changes in exams differed by discipline and even by course, apparently depending on an interplay between departmental culture, course organization, and perceived course ownership, demonstrating the complex nature of transformation in higher education. We conclude that while transformation must be supported at all organizational levels, ultimately, change is controlled by factors at the course and departmental levels.

Developing an Analytical Framework to Characterize Student Reasoning about Complex Processes

Real-world processes are complex and require ideas from multiple disciplines to be explained. However, many science courses offer limited opportunities for students to synthesize scientific ideas into coherent explanations. In this study, we investigated how students constructed causal explanations of complex phenomena to better understand the ways they approach this practice. We interviewed 12 undergraduate science majors and asked them to explain real-world phenomena. From these interviews, we developed a characterization framework that described the reasoning patterns we found. In this framework, we identified three explanatory frames that differentiated the kinds of explanations students provided: a colloquial frame, wherein participants activated conceptual resources based on personal experience using everyday language; an emerging mechanistic frame, wherein participants used scientific concepts in semicoherent ways; and a causal mechanistic frame, wherein participants cohesively drew upon scientific conceptual resources to construct mechanistic explanations. Overall, the causal mechanistic frame was the least prevalent frame invoked by students. Instead, many drew on an emerging mechanistic frame and struggled to identify and apply scientific concepts to real-world scenarios. We advocate for incorporating opportunities to reason about real-world phenomena into undergraduate science curricula to provide students with experience integrating scientific concepts to explain real-world phenomena.