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Flow Cytometry

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.
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Related Experiment Video

Updated: Jul 20, 2026

Analysis of Nonhomologous End Joining and Homologous Recombination Efficiency in HEK-293T Cells Using GFP-Based Reporter Systems

Analysis of Nonhomologous End Joining and Homologous Recombination Efficiency in HEK-293T Cells Using GFP-Based Reporter Systems

Published on: February 2, 2024

A simple analysis system for the estimation of recombination efficiency using fluorescence-activated cell sorting.

Min Soo Kim¹, Won Hee Kim, Gyun Min Lee

¹Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1 Kusong-Dong, Yusong-Gu, Daejon 305-701, Republic of Korea.

Journal of Biotechnology

|August 29, 2006

Summary

A new fluorescence-activated cell sorting (FACS) system accurately estimates in vivo recombination efficiency. Spacer mutations significantly impact recombination, with F(3) and m2 mutants showing higher efficiency than F(5) and lox2272 mutants.

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Area of Science:

Molecular Biology
Biotechnology
Genetics

Background:

Site-specific recombination is crucial for genetic engineering and gene therapy.
Accurate measurement of recombination efficiency is essential for optimizing these applications.
Existing methods for efficiency estimation can be laborious and less precise.

Purpose of the Study:

To develop and validate a simple, accurate fluorescence-activated cell sorting (FACS) based system for in vivo recombination efficiency estimation.
To compare recombination efficiencies of different spacer mutants using Flpe and Cre recombinase systems.
To investigate the impact of spacer sequence variations on recombination efficiency.

Main Methods:

Design and implementation of a novel FACS analysis system for quantifying recombination.
Utilized F(3)/F(5) and m2/lox2272 mutant sequences for Flpe- and Cre-mediated cassette exchange, respectively.
Employed PCR analysis to confirm site-specific recombination mediated by the exchange cassettes.

Main Results:

The developed FACS system accurately estimated in vivo recombination efficiency.
Spacer mutant sequences exhibited varying recombination efficiencies, with F(3) and m2 outperforming F(5) and lox2272.
Recombination efficiency is influenced by the spacer region sequence, beyond its interaction with wild-type sequences.

Conclusions:

The FACS-based system is a feasible and effective tool for estimating recombination efficiency.
Spacer sequence modifications significantly alter recombination efficiency in site-specific recombination.
This system offers a precise and efficient method for evaluating diverse mutant candidates for recombination studies.