HISTORY OF IN SITU HYBRIDIZATION

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Hi friends, 

I could not post regularly due to some minor health issues. Hope I will be regularly able to post from now on.  Thank you everyone for your constant support and suggestions. 


In this post, I will take you through the history of in situ hybridization (ISH).


What is ISH?


In situ hybridization (ISH) is a cytogenetic technique which allows high-resolution detection, quantification and localization of nucleic acid targets inside cells or tissues. The method is based on the hybridization of sequence specific complementary probes (typically DNA sequences) to their target inside the cell. Binding of complementary probes to the target of interest enables its detection and visualization. 

 

Picture taken from D.Huber et al. Micro and nano engineering (1) 2018 15-24


Historical overview on ISH


 Immunofluorescence based protein detection methods were first used by Coons et al in 1941.  

Ø  Melting and re-hybridization of DNA and complementary RNA-DNA binding were first described in the 1960s.  

Ø  In 1969, Pardue and Gall tagged rRNA probes with H3 to autoradiographically visualize rRNA encoding genes in cytological preparations of Xenopus laevis oocytes. 

Ø  Simulatneously, John et al, Buongiorno-Nardelli, Amaldi independently developed in situ hybridization techniques for the detection of rDNA in Xenopus laevis in paraffin-embedded sections. 

Ø  Few years later, Gall et al demonstrated use of ISH in mammalian cells, he visualized satellite elements in heterochromatic regions of mouse chromosomes. 


      There were several drawbacks of this radioisotopic hybridization. 


ü  These radioactive probes were unstable as the isotope decays over time and the specific activity of the probe is not constant. 

ü  Though the sensitivity of radiography is high but resolution is limited. 

ü  Long exposure times are often required to produce measurable signals on radiography film, delaying results of the assay.

ü  Radiolabeled probe is relatively costly and hazardous material and it must be transported, handled, stored and disposed of in accordance with regulations.


 Owing to these drawbacks, experiments were going on for better in situ hybridization techniques where non-isotopic labeling approaches for ISH probes were tried. 


 We shall see in the further posts how the shift from radioactive to fluorescent labeled probes came into use. 


References: 

1.D.Huber et al. Fluorescence in situ hybridization (FISH): History, limitations and what to expect from micro-scale FISH? Micro and Nano Engineering 1 (2018) 15-24. 

2. Jeffrey M Levsky and Robert H. Singer. Fluorescence in situ hybridization: past, present and future. Journal of Cell Sciences 116 (14).

3. Joseph G. Gall. The Origin of In situ Hybridization- a Personal History. Methods.2016 April 1; 98:4-9.

 

                                                                                Written by Dr.Priyavadhana B

 

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