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Fluorescence In Situ Hybridization (FISH) - Advanced Technologies for Single Cell Visualization and Identification

Research infrastructure FISH can be used to detect specific gene signatures in intact organisms – without nucleic acid extraction, PCR and gene sequencing. FISH can be evaluated by fluorescence microscopy, flow cytometry or by other methods. Some of the main benefits of FISH is that the morphology of the targeted object remains intact and that its distribution and association to others can be evaluated in the natural environment. FISH is used worldwide in medical science, life science, and biotechnology.

FISH method description

Background

Fluorescence In Situ Hybridization (FISH) was developed in the 1980s and together with PCR and gene sequencing, these methods revolutionized many fields in biology (from chromosomal diagnostics to the many fields in microbiology, ecology and biotechnology). Today, PCR, gene sequencing and omics have become the most common diagnostic methods, simply because they are easier to automate and provide precise quantitative data. However, these methods are not seldom time-consuming and bias-free, they also fail to describe relevant parameters such as morphology, distribution and association between different objects in their own environment.

Usefulness of FISH

FISH is a relatively fast and straightforward method and can thus complement the PCR and gene sequencing/omics diagnostic approach in several ways:

  1. quick identification (from 1 to 96 h, depending on target and goal) of different kinds of intact cells (Archaea, Bacteria, Eukarya) and to some extent viruses;
  2. pre-screening method prior to time-consuming techniques;
  3. follow-up method for specific questions based on a large amount of data;
  4. combination with a multitude of other methods.

Principle of FISH

In simple terms, FISH can be regarded as a phylogenetic staining method, as it is based on gene probes that are designed to target specific gene sequences. These gene probes are labeled with different kinds of markers – mostly fluorochromes. Since several different gene probes can be used simultaneously in one FISH experiment, different gene targets can therefore be easily distinguished by different colours. FISH experiments can be evaluated by different techniques, such as with microscopy and flow cytometry. 

Disadvantages of FISH

Successful FISH depends on several parameters, such as optimal penetration of gene probes through the cell wall, amount of gene copy numbers of the intended gene target, stringent and specific gene probes, and a non-fluorescent background. If these demands are not met, different kinds of biases may arise. To overcome such obstacles, a plethora of different kinds of FISH protocols have been developed.

Requirements for FISH / service suggestions

  • Appropriate fixation and storage of sample prior to FISH
  • Detailed knowledge about the phylogeny and suitability of the gene probes
  • Selection of appropriate FISH protocols
  • Selection of suitable references and controls
  • Selection of suitable detection technique
  • Performance and evaluation of FISH experiments

Evaluation options of FISH results

FISH can be evaluated in different ways and combined with other methods for optimal retrieval of additional information.

Options available at Umeå University are:

  • Fluorescence microscopy (BICU, other SciLifeLab facilities),
  • Electron microscopy (UCEM, other SciLifeLab facilities),
  • Vibrational spectroscopy (VISP),
  • Flow cytometry (Flow@CliMi)

Other facilities:

  • Stable isotope techniques (SLU or EMG)
  • Single Cell Genomics (SciLifeLab Uppsala)
  • nanoSIMS in Sweden (Gothenburg)

Poster: In situ detection and fluorescence microscopy of genes and cells in their natural environments

In situ detection and fluorescence microscopy of genes and cells in their natural environments

Current projects

Different FISH procedures are used in a wide range of projects at Umeå University and with collaborators in Sweden or in other countries, such as projects in the fields of insect microbiology, plant microbiology, geomicrobiology, astrobiology, medical microbiology, gut microbiology, food microbiology, probiotics, environmental microbiology, biological wastewater treatment, fungal biotechnology, wood biotechnology and developmental biology.

(Download FISH presentation slide as PDF)

Course activities

Planned courses related to FISH (digital and where possible live) – dates will be announced within soon

Contents:

1a-b) Introduction to FISH
2a-b) Introduction into phylogeny and gene probe design and the bioinformatics software package ARB
3a-b) Advanced FISH applications
4) Brain-storming seminar on participants' own plans for FISH experiments
5) Summary

Mode: Lectures, computer practice and lab exercises.

Course organisers:

  • Advanced FISH applications in medical sciences. Organisers: Natuschka Lee, UmU, in collaboration with the Biofilms Research Center at Interfaces, Malmö University.
  • Advanced FISH applications in environmental sciences. Organisers: Natuschka Lee, within the South Africa-Sweden University Forum, in collaboration with the North-West University in Pretoria, South Africa.

Contact

Natuschka Lee
Research fellow
E-mail
Email

Publications

Publications

Examples of publications using the FISH technics in different projects