Build Your Own Microscope: Step-By-Step Guide for Building a Prism-Based TIRF Microscope.
Dalton R GibbsAnisa KaurAnoja MegalathanKumar SapkotaSoma DhakalPublished in: Methods and protocols (2018)
Prism-based total internal reflection fluorescence (pTIRF) microscopy is one of the most widely used techniques for the single molecule analysis of a vast range of samples including biomolecules, nanostructures, and cells, to name a few. It allows for excitation of surface bound molecules/particles/quantum dots via evanescent field of a confined region of space, which is beneficial not only for single molecule detection but also for analysis of single molecule dynamics and for acquiring kinetics data. However, there is neither a commercial microscope available for purchase nor a detailed guide dedicated for building this microscope. Thus far, pTIRF microscopes are custom-built with the use of a commercially available inverted microscope, which requires high level of expertise in selecting and handling sophisticated instrument-parts. To directly address this technology gap, here we describe a step-by-step guide on how to build and characterize a pTIRF microscope for in vitro single-molecule imaging, nanostructure analysis and other life sciences research.
Keyphrases
- single molecule
- atomic force microscopy
- living cells
- quantum dots
- induced apoptosis
- high resolution
- cell cycle arrest
- big data
- electronic health record
- photodynamic therapy
- sensitive detection
- loop mediated isothermal amplification
- signaling pathway
- single cell
- fluorescence imaging
- data analysis
- label free
- real time pcr
- optical coherence tomography