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Welcome to the blog
The content posted here is intended to help and is meant to be as accurate as possible. There will inevitably be mistakes or gaps in the knowledge. This is not peer-reviewed science, but is evidence-based as much as possible. If you see a factual inaccuracy or disagree, send in a correction to colibri.cytometry at gmail. Also, while some of this content tries to push the boundaries of what we know about how to do flow, it is not intended to claim invention credit. If you feel a reference has been overlooked, please let me know.
Commenting has been deactivated. Sorry.
The comments were getting spammed.
Also, GDPR is a headache.
Notes on the dish soap protocol
This protocol is good for when you are trying to combine staining for antigens in different cellular compartments. It is not the single best solution for any one thing.
Dec 16, 20258 min read
AutoSpectral: Full workflow example
A full workflow example for using AutoSpectral to perform spectral unmixing
Nov 24, 20258 min read
AutoSpectral: ID7000 40C examples
Today, let's look at some 40-colour data from a 5-laser Sony ID7000, using AutoSpectral for the unmixing.
Oct 30, 20252 min read
AutoSpectral: OMIP-102 examples
What happens if we run AutoSpectral on a good dataset, with cell-based controls?
Oct 29, 20252 min read
Introducing AutoSpectral: an optimized unmixing workflow
With AutoSpectral, we provide an optimized, complete pipeline in R to make the best use of the information in the single-stained controls and reproducibly generate better unmixing.
Oct 28, 20253 min read
AutoSpectral: Per Cell Fluorophore Optimization
The aim of this article is to explain a bit more about how we go about optimizing the fluorophore signatures used for unmixing on a per-cell basis in AutoSpectral. Let me know if the explanation doesn't make sense.
Oct 27, 20259 min read
AutoSpectral: Single Cell Autofluorescence
The aim of this article is to show you how to use AutoSpectral’s per-cell autofluorescence (AF) extraction. This method selects the best AF signature to use for each cell (or point of debris, noise, whatever) in the data.
Oct 27, 20255 min read
AutoSpectral: plotting
In this article, let’s look at a couple of the functions that AutoSpectral offers for plotting your spectra and data. AutoSpectral isn't intended to replace flow analysis software like FlowJo or FCS Express, but it's essential to have some options for visualizing the outputs.
Oct 27, 20254 min read
AutoSpectral: reopening and rerunning
In this article, we'll cover some shortcuts to allow you to return to AutoSpectral once you've already run an experiment.
Oct 27, 20254 min read
AutoSpectral: cleaning
In this article, we’ll try to cover the functions that AutoSpectral offers for cleaning your single-stained control data.
Oct 27, 20257 min read
AutoSpectral: gating
In this article, we’ll try to cover some of the features and tuning for the automated gating in AutoSpectral.
Oct 27, 20257 min read
AutoSpectral: creating the control file
In this article, we’re going to cover how to create a control file, which is an essential first step in running AutoSpectral. The control file is a specifically formatted spreadsheet describing your single-stained control files.
Oct 27, 20255 min read
AutoSpectral: basic workflow
In this article, I'll cover the basic workflow for using AutoSpectral for unmixing spectral flow cytometry data.
Oct 27, 20252 min read
Spectral mixing matrix normalization
Should we normalize the fluorophore signatures to the peak or to the sum of all the signals?
Sep 22, 20253 min read
Why do we get autofluorescence problems?
Today the aim is to illustrate visually how cellular autofluorescence ends up in our unmixed flow cytometry data, confounding our results.
Sep 8, 20252 min read
What's an unmixing matrix?
In the last post, we looked at what the spectral mixing (sometimes called spillover) matrix looked like. Today, let's go through a bit about the unmixing matrix.
Aug 26, 20255 min read
Cookies...
In order to avoid any issues with GDPR or other internet laws, I'm taking a couple of less pleasant steps. First, you'll now see the...
Aug 24, 20251 min read
What's in a spectral matrix?
Today the plan is to try to explain what a spillover or spectral matrix is
Aug 11, 20257 min read
AutoSpill 2: Spectral Boogaloo
AutoSpectral is a new way of getting better unmixing.
Jun 30, 20251 min read
Stabilizing your unmixing: drift
Today's post is meant to illustrate why I do not recommend running new samples using the same unmixing matrix across multiple days.
May 12, 20252 min read
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