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Light, Imaging, Vision: An Interdisciplinary Undergraduate Course
Tuesday, February 14, 2017 2275-Pos Board B595 A Ratiometric Method to Measure Viscosity Inside Mesoporous Silica Particles using Protein-Bound Fluorescent Probe ˚ kerman. Pegah S. Nabavi Zadeh, Bjo¨rn A Chemistry and Chemical Engineering, Chalmers, Go¨teborg, Sweden. Mesoporous silica particles are used for immobilization of enzymes in order to increase enzyme stability, facilitate product purification and reuse of enzyme. Many efforts have been done to investigate how the environment inside the pores changes after enzyme immobilization and how it can affect activity and stability of immobilized enzymes. One of these environmental changes can be viscosity inside the pores and comparing the difference with the bulk solution. Here, we investigate viscosity inside mesoporous silica particles, SBA-15, 1mm diameter with 9nm pore diameter, based on two different cyanine fluorophores which are sensitive to viscosity, Cy3 and Cy5 using a ratiometric method. The probes can bond covalently to lipase as enzyme used in two different conditions: a. both probes seated on one enzyme, b. they can be attached to the enzyme separately. We work with a ratiometric method to measure only viscosity and no other solvent effects. In this study, Cy5 can be used to normalize for polarity other than viscosity. in addition, we take advantage of the energy transfer (FRET) between probes when they are seated on one enzyme and measure the distance between them. 2276-Pos Board B596 Making a Puzzle Microscope as a Strategy to Recruit High School Girls and Minorities in Biophysics Yuly E. Sanchez. Universidad Nacional de Colombia, Bogota, Colombia. Low participation of woman in science is an actual fact that has been discussed by different sources, women’s organizations, prestigious universities and different journals. According to UNESCO ‘‘the world needs science and science needs women’’, that’s why last February 11th 2016 was established as the first International Day of Women and Girls in Science, UNESCO’s message was clear - the new Agenda will not meet its promise without investing in women’s and girls’ empowerment through and in science. Unfortunately women in Colombia have been victims of both armed conflict and a society run by men and her role in scientific areas is not increasing. According to recent research conducted at the National University of Colombia, women have little participation in university courses related to the exact sciences, which is easily verified with statistics about of women’s participation in science careers as technology, engineering and mathematics (STEM). As a contribution to address this topic, we proposed an activity aimed at recruiting high school girls in biophysics and science in general. Our proposal was to build a puzzle wooden microscope kit by Echo LaboratoriesÒ, this activities where performed with 4 different groups of 25-30 girls and 3 groups of 25-30 students from minorities as afro-Colombian and older students, as a complement we explored different tissues with our microscope and we compared it with a conventional school microscope. Is well known that puzzles are also an important educational learning tool for young children as they provide many skills and mental learning benefits and opportunities, we used this concept too for elderly students. We found that after they build the wooden scopes, they were able to conclude that was much like a science experiment. This proposal incorporating this microscope as educational puzzles into today’s school curriculum is a great way to encourage their participation in science. 2277-Pos Board B597 Combining Molecular Visualization with Bench Methods in a HypothesisDriven Undergraduate Biochemistry Lab Course Thomas Holt1, Rebecca Roberts1, Julia R. Koeppe2, Paul A. Craig3. 1 Ursinus College, Collegeville, PA, USA, 2Chemistry, SUNY Oswego, Oswego, NY, USA, 3Rochester Institute of Technology, Rochester, NY, USA. We are developing an undergraduate biochemistry lab curriculum based on authentic inquiry. Students on six campuses are combining computational (in silico) and wet lab (in vitro) techniques as they characterize enzymes whose three dimensional structures are known but to which functions have not been previously ascribed. The in silico modules include protein visualization with PyMOL, structural alignment using Dali and ProMOL, sequence exploration with BLAST and Pfam, and ligand docking with PyRX and Autodock Vina. The goal is to predict the function of the enzymes and to identify promising substrates for the active sites. In the wet lab, students express and purify the target enzymes and perform kinetic assays with substrates selected from their docking studies. Their learning as students and their growth as scientists is being assessed in terms of research methods, visualization, biolog-
463a
ical context, and mechanism of protein function. The lab course is an extension of successful undergraduate research efforts at RIT and Dowling College. The modules that are developed have been disseminated to the scientific community via a web site (promol.org), including both protocols and captioned video instructions for the techniques involved. Over the course of the project, we have followed changes in faculty and teaching assistant competence in two areas: effective teaching with both in silico and in vitro tools and the development of skills in the area of measuring learning gains by students. We are also analyzing the flexibility of the curriculum for adoption as a whole in a single course or in parts across multiple courses. As we conduct the lab on the different campuses, we are focusing on advantages of our approach and barriers to implementation that exist on each campus., from the level of student acceptance and faculty training, to resources that are needed, to changes in the culture at the departmental and institutional levels. We are now optimizing the curriculum for wider dissemination, and we seek input from additional potential adopters about their level of interest and the barriers that they anticipate on their campuses. This project is supported in part by NSF IUSE 1503699. 2278-Pos Board B598 Expanding the Scope of Single Molecule FRET Spectroscopy towards Primarily Undergraduate Institutions Jesse Howe, Gregory Walters, Kambiz Hamadani. CSU San Marcos, Vista, CA, USA. Single molecule fluorescence methods provide a unique and ultrasensitive set of probes for monitoring the conformations and dynamics of protein chains within physiologically relevant contexts. In particular, single molecule fluorescence resonance energy transfer (smFRET) is capable of monitoring the distances between two fluorescent dyes with sub-nanometer spatial and nanosecond temporal resolution. Unfortunately, smFRET remains inaccessible to many biophysical researchers and most of all to students at primarily undergraduate research institutions. This is in part due to the difficulties involved in making dual-labelled protein samples but also stems from the expense and/or complexity of existing single-molecule fluorescence detection platforms. To address the first issue, we have developed a sample generation protocol suitable for making large libraries of dual-labelled proteins and ribosome-bound nascent chains (RNCs). This method consists of using a purified and reconstituted in-vitro translation system for protein expression, the incorporation of azide or alkyne-bearing non-standard amino acids, and finally clickchemistry-based dye attachment. Here, we show that this approach is simple and robust enough to be used by undergraduate students to make large libraries of dual-labelled protein samples in about a day. To enable smFRET-based screening of these libraries we have also built the first-ever confocal smFRET microscope at a primarily undergraduate institution. This instrument was built for less than $20K largely using second-hand optical components. It has a 3-axis piezo-scanning stage which enables single-molecule imaging and trajectory analysis on surface-immobilized molecules. It also has alternating laser excitation capabilities which enable digital sorting of single molecules using multi-dimensional histograms. This work marks the first implementation of smFRET as a research tool at a primarily undergraduate research institution. 2279-Pos Board B599 Light, Imaging, Vision: An Interdisciplinary Undergraduate Course Philip Nelson. Physics/Astronomy, Univ Pennsylvania, Philadelphia, PA, USA. Students in physical and life science, and in engineering, need to know about the physics and biology of light. In the twenty-first century, it has become increasingly clear that the quantum nature of light is essential both for the latest imaging modalities and even to advance our knowledge of fundamental life processes, such as photosynthesis and human vision. But many optics courses remain rooted in classical physics, with photons as an afterthought. I’ll describe a new undergraduate course, for students in several science and engineering majors, that takes students from the rudiments of probability theory to modern experimental methods like fluorescence imaging and Fo¨rster resonance energy transfer. After a digression into color vision, students then see how the Feynman principle explains the apparently wavelike phenomena associated to light, including applications like diffraction limit, subdiffraction imaging, total internal reflection and TIRF microscopy. Then we see how scientists documented the single-quantum sensitivity of the eye seven decades earlier than ‘ought’ to have been possible, and finally close with the remarkable signaling cascade that delivers such outstanding performance. A new textbook, to be published in April 2017, allows others to replicate this course. Partially supported by the United States National Science Foundation under Grant PHY-1601894.
463a
ical context, and mechanism of protein function. The lab course is an extension of successful undergraduate research efforts at RIT and Dowling College. The modules that are developed have been disseminated to the scientific community via a web site (promol.org), including both protocols and captioned video instructions for the techniques involved. Over the course of the project, we have followed changes in faculty and teaching assistant competence in two areas: effective teaching with both in silico and in vitro tools and the development of skills in the area of measuring learning gains by students. We are also analyzing the flexibility of the curriculum for adoption as a whole in a single course or in parts across multiple courses. As we conduct the lab on the different campuses, we are focusing on advantages of our approach and barriers to implementation that exist on each campus., from the level of student acceptance and faculty training, to resources that are needed, to changes in the culture at the departmental and institutional levels. We are now optimizing the curriculum for wider dissemination, and we seek input from additional potential adopters about their level of interest and the barriers that they anticipate on their campuses. This project is supported in part by NSF IUSE 1503699. 2278-Pos Board B598 Expanding the Scope of Single Molecule FRET Spectroscopy towards Primarily Undergraduate Institutions Jesse Howe, Gregory Walters, Kambiz Hamadani. CSU San Marcos, Vista, CA, USA. Single molecule fluorescence methods provide a unique and ultrasensitive set of probes for monitoring the conformations and dynamics of protein chains within physiologically relevant contexts. In particular, single molecule fluorescence resonance energy transfer (smFRET) is capable of monitoring the distances between two fluorescent dyes with sub-nanometer spatial and nanosecond temporal resolution. Unfortunately, smFRET remains inaccessible to many biophysical researchers and most of all to students at primarily undergraduate research institutions. This is in part due to the difficulties involved in making dual-labelled protein samples but also stems from the expense and/or complexity of existing single-molecule fluorescence detection platforms. To address the first issue, we have developed a sample generation protocol suitable for making large libraries of dual-labelled proteins and ribosome-bound nascent chains (RNCs). This method consists of using a purified and reconstituted in-vitro translation system for protein expression, the incorporation of azide or alkyne-bearing non-standard amino acids, and finally clickchemistry-based dye attachment. Here, we show that this approach is simple and robust enough to be used by undergraduate students to make large libraries of dual-labelled protein samples in about a day. To enable smFRET-based screening of these libraries we have also built the first-ever confocal smFRET microscope at a primarily undergraduate institution. This instrument was built for less than $20K largely using second-hand optical components. It has a 3-axis piezo-scanning stage which enables single-molecule imaging and trajectory analysis on surface-immobilized molecules. It also has alternating laser excitation capabilities which enable digital sorting of single molecules using multi-dimensional histograms. This work marks the first implementation of smFRET as a research tool at a primarily undergraduate research institution. 2279-Pos Board B599 Light, Imaging, Vision: An Interdisciplinary Undergraduate Course Philip Nelson. Physics/Astronomy, Univ Pennsylvania, Philadelphia, PA, USA. Students in physical and life science, and in engineering, need to know about the physics and biology of light. In the twenty-first century, it has become increasingly clear that the quantum nature of light is essential both for the latest imaging modalities and even to advance our knowledge of fundamental life processes, such as photosynthesis and human vision. But many optics courses remain rooted in classical physics, with photons as an afterthought. I’ll describe a new undergraduate course, for students in several science and engineering majors, that takes students from the rudiments of probability theory to modern experimental methods like fluorescence imaging and Fo¨rster resonance energy transfer. After a digression into color vision, students then see how the Feynman principle explains the apparently wavelike phenomena associated to light, including applications like diffraction limit, subdiffraction imaging, total internal reflection and TIRF microscopy. Then we see how scientists documented the single-quantum sensitivity of the eye seven decades earlier than ‘ought’ to have been possible, and finally close with the remarkable signaling cascade that delivers such outstanding performance. A new textbook, to be published in April 2017, allows others to replicate this course. Partially supported by the United States National Science Foundation under Grant PHY-1601894.