Calcium phosphate as a gene carrier: electron microscopy.
Transfection of DNA into cultured cells by the calcium phosphate (CaPi)-mediated method provides a means of examining gene functions. Several factors in preparing the CaPi-DNA co-precipitates were speculated to be correlated with the transfection efficiency. To achieve a better understanding of the steps of CaPi formation and the effect of other components in the medium, this study was undertaken to examine the morphological changes of CaPi in the transfection medium. Transmission electron microscopy (TEM) was employed to investigate the reaction products of CaPi precipitates prepared in the presence or absence of plasmid DNA, Dulbecco's modified Eagle medium and calf serum. The effect of pH of the HEPES-buffered saline (HBS) in which CaPi was formed was also examined. Here we report the observations we obtained from TEM. The results showed that CaPi precipitates underwent several steps of conversion in the presence of other components in the cell culture medium. The pH of the HBS in which CaPi precipitates were formed also exerts a profound effect on the morphology of the CaPi solid phase, indicating that the associated compositional and structural changes occurred during the maturation of CaPi in the DNA transfection medium.
Yang
Y W
YW
School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan.
Yang
J C
JC
D002130:Calcium Phosphates / Q000737:chemistry*; D002478:Cells, Cultured; D003102:Colloids; D003470:Culture Media; D006863:Hydrogen-Ion Concentration; D008854:Microscopy, Electron; D010957:Plasmids / Q000737:chemistry / Q000648:ultrastructure; D014162:Transfection
D002130:Calcium Phosphates; D003102:Colloids; D003470:Culture Media