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中国生物工程杂志

China Biotechnology
China Biotechnology  2009, Vol. 29 Issue (07): 62-67    DOI:
    
Effect of Paraformaldehyde Fixation on the FRET Efficiency Calculation between Fluorescent Proteins
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Abstract  Abstract Objective: To investigate the effect of paraformaldehyde fixation on measuring the protein-protein interaction by fluorescence resonance energy transfer (FRET) to resolve the problem of FRET efficiency calculation in excess-movement cells. Methods: The C terminals of TCR α chain (TRA) and TCR β chain (TRB) genes, which were ideal for protein-protein interaction research, were fused with ECFP and EYFP gene respectively by fusion PCR and transferred into target cell. A grou Pcells were fixed in paraformaldehyde (0.5%) for 0.5~1h and another left alive, then these cells were subject to ECFP/EYFP FRET calculation with confocal laser scanning microscope. The ECFP/EYFP FRET efficiencies in live and fixed cell were analyzed and compared. Results: There is no significant statistical difference between the ECFP/EYFP FRET efficiencies of live cell and cell fixed with lower paraformaldehyde concentration and shorter incubation time. Conclusion: fixation with low-concentration paraformaldehyde and short-time incubation has no distinct influence on measuring protein-protein interaction, and facilitated the FRET calculation in excess-movement cells.

Key wordsFixation;Paraformaldehyde;FRET;ECFP/EYFP     
Received: 08 January 2009      Published: 28 July 2009
ZTFLH:  中图分类号Q331  
Corresponding Authors: SHAO Hong-Wei     E-mail: shulhuang@sina.com
Cite this article:

SHAO Gong-Wei- Zhang-Wen-Feng- Hu-Jing-Lian- Chen-Han- Tun-Feng-Lin- Huang-Shu-Lin. Effect of Paraformaldehyde Fixation on the FRET Efficiency Calculation between Fluorescent Proteins. China Biotechnology, 2009, 29(07): 62-67.

URL:

https://manu60.magtech.com.cn/biotech/     OR     https://manu60.magtech.com.cn/biotech/Y2009/V29/I07/62

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