Microscopes are very complex pieces of equipment, that have evolved to accommodate many different imaging techniques. Microscopy and its applications have promoted many key breakthroughs in life sciences. Its impact can be seen by the numerous nobel prizes in Physics, Chemistry and Physiology and Medicine, that have been attributed to the development of new microscopy applications and techniques, or from discoveries that were possible due to microscopy breakthroughs.
At the Andor Microscopy School, we aim to give life science researchers an overview of the microscopy world, from the fundamental applications to more recent high-end techniques. We will start with an introduction to microscopy, its history and simultaneously introduce concepts of optics that are important to understand how microscopes work.
This free course will also introduce concepts in detectors, fluorescence microscopy, sample preparation and confocal microscopy. Once the fundamental keystones of the course are laid, we will introduce advanced microscopy applications such as TIRF, super-resolution and photostimulation techniques. Finally, we will present an introduction to post-processing and image analysis tools.
The Andor Microscopy School will run on Wednesdays. This is a free course designed for researchers who aim to improve their microscopy knowledge. Do you want to enhance your understanding of microscopy? Then you should register for the Andor Microscopy School.
Lesson 1 – The History of Microscopy
This lesson gives an overview of the history of microscopy and simultaneously introduces some concepts in optics that are important for understanding microscopes, its components how it affects the resulting image.
Lesson 2 – Transmitted Light Microscopy
This lesson reviews the anatomy of the microscope and essential concepts in microscopy, such as numerical aperture and resolution. We will explain the bases of the differences between the different transmitted light techniques. Finally, to give a clear understanding between the optical requirement for the techniques and the resulting image, we will show images acquired with the different techniques.
Lesson 3 – Microscopy Cameras - Fundamentals of Digital Imaging and Sensor Technologies
In this first of two lessons on microscopy cameras we cover the fundamentals of microscopy cameras. We explore what are the key parameters in images we take on a microscope from a camera’s perspective. Then we break down the anatomy of a modern camera. Finally, we look at the different sensor technologies that have been developed, leading up to the current technologies in use today. We address some of the common questions such as how cameras work, why would you use a mono camera or a colour camera for imaging and what does back-illumination mean? This sets the background for the following lesson that compares the different camera technologies and how they suit different microscopy applications.
Lesson 4 - Microscopy Cameras - Comparing Camera Technologies and Matching them to Applications
In this second lesson of two modules on the topic of imaging cameras we explore how the different camera technologies compare relative to the parameters covered in the first module (Lesson 3 – Microscopy Cameras - Fundamentals of Digital Imaging and Sensor Technologies). We discuss the performance of these camera technologies available for sensitivity, field of view and speed. Several applications are summarized against their main imaging requirements. Suitable camera types are then suggested for these based on their ability to fulfil these criteria
Lesson 5 - Principles of Fluorescence Light Microscopy
In this lesson we present the concept of fluorescence, and how is this physical propriety is applied to microscopy. We will give a brief overview of the hardware required for a fluorescent microscope, as well as discuss some concerns that should be considered when starting a fluorescence imaging protocol. Overall, the goal is that individuals that attend this lesson gain/refresh their knowledge about fluorescence microscopy potential. We hope to arm researchers with better tools to plan their work.
Lesson 6 - Principles in Confocal Microscopy
There are two types of confocal microscopes - single pinhole confocal, or point scanners, where a single pinhole will discard the out of focus light and multiple point confocal, or spinning disks, where multiple pinholes will discard the out of focus light. In this lesson we explain what a confocal microscope is and discuss the differences between single-point confocal and multipoint confocal. By the end of this talk, we hope that you will have a clear understanding of confocal microscopy and it´s applications in life sciences.
Lesson 7 - Sample Preparation for Immunohistochemistry
In this lesson we discuss every aspect of sample preparation for Immunohistochemistry. There are several steps for Immunohistochemistry sample preparation. It begins with adequate sample collection and fixation. Subsequently, the samples are stained with antibodies. Antibody staining is crucial and dependent on several factors that must be determined experimentally. In recent years together with the increasing microscopy power to image large samples, several post-treatment clearing techniques have emerged, allowing in toto imaging of fluorescently labelled samples. Currently, all these new technological advances enable the imaging of biological samples with an unprecedented morphological contextualization.
Lesson 8 - Lifting the Cellular Fog with Total Internal Reflection Fluorescence (TIRF) Microscopy
This microscopy course lecture covers the basics of TIRF microscopy, the necessities that make for a successful TIRF imaging experiment, the challenges of TIRF image analysis, and discusses how TIRF microscopy in combination with other imaging modalities – either simultaneously or sequentially – have advanced our understanding of complex cellular processes and molecular dynamics. Some of the latest developments of this technique will also be reviewed.
Lesson 9 and 10 - Super Resolution Microscopy
In this microscopy lesson, we cover the basics of how the three most common super-resolution techniques (SIM, STED and SMLM) work, including factors such as labelling, hardware, image processing, and the advantages and disadvantages of the methods. We will also see examples of how different super-resolution techniques have been used to successfully address different biological problems. Finally, we will examine the current state-of-the art and future directions for the field of super-resolution microscopy.
Lesson 11 - Principles of Deconvolution
In this microscopy course lesson we focus on optical microscopy and outline concepts of image formation by convolution; microscope point spread and optical transfer functions; and how these can be applied to develop simple and iterative deconvolution algorithms. Dr Browne will present examples and identify common pitfalls to be avoided.