Imaging Instrumentation Considerations For Your Immunofluorescence Experiments
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The results of your immunofluorescence (IF) experiment are impacted by several factors, including which type of microscope and imaging system you choose. In this edition of Learning Lab LIVE!, IF Expert Virginia Bain, PhD, discusses what you should keep in mind when starting an IF experiment.
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Table of Contents:
0:00 Introduction
0:40 Overview
02:36 Magnification considerations
3:46 Widefield epifluorescence basics
4:47 Image capture in widefield
5:40 Widefield imaging settings
7:40 Confocal microscopy basics
9:22 Confocal imaging settings
12:33 Section thickness and
13:34 When to consider using z-stacks
14:22 Using validated reagents or validation yourself
16:33 Sample quality
17:46 What are the limitations of epifluorescenct vs. confocal microscopy?
18:58 How can I deal with high background in parrafin-embedded tissues when performing immunofluorescence?
20:43 Any tips for immunofluorescence combined with in situ hybridization?
22:25 How do different blocking buffers compare of IF?
24:24 What instrument is best for imaging protein localization?
25:54 Can I image whole brain with confocal?
27:19 What fixation is best for brain tissue? Is it better to use frozen?
28:23 What's the difference between and inverted and upright microscopes?
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