Precise Identification of Gene Products in Hearts after in Vivo Gene Transfection, Using Sendai Virus-Coated Proteoliposomes

• Published in: Biochemical and biophysical research communications Vol. 259; no. 2; pp. 408 - 413
• Main Authors: Kawada, Tomie, Shin, Wee Soo, Nakatsuru, Yoko, Koizumi, Toshiyuki, Sakamoto, Aiji, Nakajima, Toshiyuki, Okai-Matsuo, Yoko, Nakazawa, Mikio, Sato, Hiroshi, Ishikawa, Takatoshi, Toyo-Oka, Teruhiko
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 07-06-1999
• Subjects: Animals; beta-Galactosidase - analysis; beta-Galactosidase - genetics; cardiomyocytes; gene transfection in vivo; Genes, Reporter; green fluorescence protein; Green Fluorescent Proteins; Heart - virology; Histocytochemistry; Luminescent Proteins - analysis; Luminescent Proteins - genetics; Male; Microscopy, Fluorescence; Myocardium - metabolism; Proteolipids - genetics; Rats; Rats, Wistar; Respirovirus - genetics; Sendai virus; Sendai virus-coated proteoliposomes; Transfection - methods; β-galactosidase; cardiomyocytes; β-galactosidase; Sendai virus-coated proteoliposomes; green fluorescence protein; gene transfection in vivo
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.1999.0764

Both efficient gene transfer and the exact identification of gene product are required for gene therapy. Gene transfection of green fluorescence protein (GFP) might be useful for the reporter. After in vivo cotransfection of GFP and β-galactosidase (β-Gal) genes in Sendai virus-coated proteoliposomes to rat hearts, we compared the sensitivity and specificity of three methods: GFP detection, histochemical staining (HC) of β-Gal activity, and immunostaining (IS) of the β-Gal protein. Fluorescence microscopy and double staining of HC and IS revealed that both GFP and IS were equally sensitive and fourfold superior to HC at the peak of gene expression. However, different from skeletal muscle, the GFP of transfected cardiomyocytes showed two demerits: the fluorescence quenching due to the intense staining of β-Gal activity, and nonspecific autofluorescence from myocardium. Thus, specific IS would be so far the most reliable to identify the gene product in heart.