Related Experiment Video

Updated: Jan 23, 2026

Phospho Flow Cytometry with Fluorescent Cell Barcoding for Single Cell Signaling Analysis and Biomarker Discovery
08:38

Phospho Flow Cytometry with Fluorescent Cell Barcoding for Single Cell Signaling Analysis and Biomarker Discovery

Published on: October 4, 2018

21.8K

Development, application and computational analysis of high-dimensional fluorescent antibody panels for single-cell

Jolanda Brummelman1, Claudia Haftmann2, Nicolás Gonzalo Núñez2

  • 1Laboratory of Translational Immunology, Humanitas Clinical and Research Center, Rozzano, Milan, Italy.

Nature Protocols
|June 5, 2019
PubMed

Insights

This study simplifies 28-color flow cytometry panel design by providing a reference map for fluorescence spreading errors. This advance accelerates high-dimensional single-cell analysis, particularly for immune system characterization.

Area of Science:

  • Immunology
  • Cell Biology
  • Biotechnology

Background:

  • Single-cell analysis is revolutionizing biological research, especially immunology.
  • Flow cytometry offers high-throughput single-cell analysis and has been extended to 28 colors.
  • Limitations of flow cytometry include autofluorescence and spreading error (SE), hindering dim marker detection and panel design.

Purpose of the Study:

  • To describe the steps for achieving 28-color flow cytometry measurement capability.
  • To provide a reference map of fluorescence spreading errors in a 28-color space.
  • To offer detailed computational analysis instructions for complex flow cytometry data.

Main Methods:

  • Development of a 28-color antibody panel.
  • Creation of a fluorescence spreading error reference map.
  • Application of computational algorithms (PhenoGraph, FlowSOM) for data analysis.

Main Results:

  • Successful implementation of 28-color flow cytometry.
  • A comprehensive reference map simplifying panel design and predicting antibody combination success.
  • Detailed computational analysis protocols for high-dimensional data.

Conclusions:

  • The described approach facilitates the design and analysis of complex, high-dimensional flow cytometry panels.
  • This method streamlines the characterization of the immune system and other biological systems.
  • The protocol is efficient, taking only a few days to complete.

More Related Videos

Analysis of Cell Suspensions Isolated from Solid Tissues by Spectral Flow Cytometry11:08

Analysis of Cell Suspensions Isolated from Solid Tissues by Spectral Flow Cytometry

Published on: May 5, 2017

13.5K
High-throughput Flow Cytometry Cell-based Assay to Detect Antibodies to N-Methyl-D-aspartate Receptor or Dopamine-2 Receptor in Human Serum10:19

High-throughput Flow Cytometry Cell-based Assay to Detect Antibodies to N-Methyl-D-aspartate Receptor or Dopamine-2 Receptor in Human Serum

Published on: November 23, 2013

16.8K

Related Concept Videos

Dimensional Analysis03:40

Dimensional Analysis

Dimensional analysis, also known as the factor label method, is a versatile approach for mathematical operations. The main principle behind this approach is: the units of quantities must be subjected to the same mathematical operations as their associated numbers. This method can be applied to computations ranging from simple unit conversions to more complex and multi-step calculations involving several different quantities and their units.
Conversion Factors and Dimensional Analysis
The unit...
60.5K
Dimensional Analysis01:27

Dimensional Analysis

Dimensional analysis is a valuable technique in fluid mechanics for simplifying complex problems by reducing them into dimensionless groups. These groups capture the essential relationships between the variables involved, allowing researchers and engineers to analyze fluid flow without dealing with each variable individually. This approach reduces the number of independent variables, allowing for easier analysis and better understanding of physical phenomena.
In fluid mechanics, dimensional...
653
Dimensional Analysis01:23

Dimensional Analysis

Dimensional analysis is a powerful tool that is used in physics and engineering to understand and predict the behavior of physical systems. The basic idea behind dimensional analysis is to express physical quantities in terms of fundamental dimensions such as the mass, length, and time. Derived dimensions like the velocity, acceleration, and force are derived from the combinations of these fundamental dimensions.
Dimensional analysis allows us to analyze and compare physical quantities on a...
2.1K
Dimensional Analysis02:19

Dimensional Analysis

The concept of dimension is important because every mathematical equation linking physical quantities must be dimensionally consistent, implying that mathematical equations must meet the following two rules. The first rule is that, in an equation, the expressions on each side of the equal sign must have the same dimensions. This is fairly intuitive since we can only add or subtract quantities of the same type (dimension). The second rule states that, in an equation, the arguments of any of the...
23.6K
Flow Cytometry01:23

Flow Cytometry

The development of flow cytometry techniques began in 1934 with initial attempts by Andrew Moldavan, a bacteriologist who counted the cells in a flowing capillary system. Moldavan pumped cells through a capillary tube focused under a microscope for visualization. The invention of photometry allowed the measurement of differentially-stained cells, and Louis Kamentsky developed the first multiparameter flow cytometer in 1965 to identify and count the cancer cells in cervical tissue specimens.
In...
15.9K
Wood Panel Products01:18

Wood Panel Products

Wood panel products are essential materials used in construction for applications such as flooring, siding, and roofing, typically available in standard dimensions of 4 feet by 8 feet, with thicknesses varying from one-quarter of an inch to one and one-eighth inches. Among the most common types of wood panels is plywood, which is produced by gluing multiple layers of thin wood veneers under pressure. The grain of the outer veneers runs lengthwise, while the grains of the interior layers run...
365