Overview # There are four steps to this process. First is to create a dataset file that allows BigSticher to interpret the data. Second is to run a macro that will flip the opposing views, so they are roughly aligned. Third is to find the best parameters that register the views together. Forth is to register and export the reconstructed data. Inital Setup # First we must install ImageJ and the BigStitcher plugin. ...
When the sample moves quickly through the field of view and you want to measure movement within the sample, digital stabilization is needed. This post will walk through how this is done using BigSticher.
For a recent blog post, I needed to get my hands on some Imaris IMS files. Fortunately for me, the good folks over at OME have a nice repository of example images in a variety of formats. The OME Team use this data for testing the read/write functions in their Bio-Formats library. I was particularly interested in downloading all of their Imaris images, but it appeared the only way to get the entire dataset was to manually download each file. I can’t waste that much time downloading files!
How to set up and use a MATLAB script to pull out ROIs in Lattice Light-sheet data.
In my previous post, I mentioned that the AIC is exploring alternative file formats for use in our processing and analysis pipelines. Our pre-commercial microscopes present a unique challenge in that they all produce different formats and structures of image data. The exact format of the output images depends on the acquisition mode of the microscope and the developer of the microscope’s control software. We are currently converting most of our images into sequences of KLB files.
There has been a lot of talk about image file formats these days. The OME team wrote a blog about it, people on the image.sc forum are discussing it, and funding groups (e.g. CZI) are putting money behind it. The AIC is certainly no stranger to the difficulties of old, outdated, and obscure image file formats. We put a fair amount of coding and computing effort into re-saving images from one format to another.
Starting in August 2019, the Advanced Imaging Center (AIC) introduced its first pre-commercial electron microscope. This system is based on work done in Janelia’s Hess lab and makes use of a powerful technique: Focused Ion Beam Scanning Electron Microscopy (FIB-SEM). This is the first in a series of blog posts that introduces this unique methodology, and the various considerations that should be made, to prospective AIC users.
A molecule will emit fluorescent light when its electron, falling from the excited state to the ground state, releases most of the energy that it absorbs during excitation. Upon returning to the ground state, the electron is ready to be excited again for another productive cycle. However, these cycles will not continue forever.
With the recent advent of several microscopy technologies, especially light sheet microscopy, with which millions of voxels can be acquired in a matter of seconds, the era of big data of imaging has arrived. The challenges in handling AIC data can be categorized into three sections: (i) data transfer, (ii) data storage, and (iii) data computation for image analysis. In this post, we discuss each section and outline some things to consider.
In a previous blog post, we discussed probe selection for single molecule localization microscopy (SMLM) techniques such as PALM and STORM. This blog post continues earlier discussions to give some practical guidance toward both probe selection and imaging considerations when performing SMLM across multiple channels.