Detection of silver nanoparticles in cells by flow cytometry using light scatter and far-red fluorescence.
The cellular uptake of different sized silver nanoparticles (AgNP) (10, 50, and 75 nm) coated with polyvinylpyrrolidone (PVP) or citrate on a human derived retinal pigment epithelial cell line (ARPE-19) was detected by flow cytometry following 24-h incubation of the cells with AgNP. A dose dependent increase of side scatter and far red fluorescence was observed with both PVP and citrate-coated 50 nm or 75 nm silver particles. Using five different flow cytometers, a far red fluorescence signal in the 700-800 nm range increased as much as 100 times background as a ratio comparing the intensity measurements of treated sample and controls. The citrate-coated silver nanoparticles (AgNP) revealed slightly more side scatter and far red fluorescence than did the PVP coated silver nanoparticles. This increased far red fluorescence signal was observed with 50 and 75 nm particles, but not with 10 nm particles. Morphological evaluation by dark field microscopy showed silver particles (50 and 75 nm) clumped and concentrated around the nucleus. One possible hypothesis to explain the emission of far red fluorescence from cells incubated with silver nanoparticles is that the silver nanoparticles inside cells agglomerate into small nano clusters that form surface plasmon resonance which interacts with laser light to emit a strong far red fluorescence signal. The results demonstrate that two different parameters (side scatter and far red fluorescence) on standard flow cytometers can be used to detect and observe metallic nanoparticles inside cells. The strength of the far red fluorescence suggests that it may be particularly useful for applications that require high sensitivity. © Published 2013 Wiley-Periodicals, Inc.
U.S. Environmental Protection Agency, Office of Research and Development, National Health and Environmental Effects Research Laboratory, Toxicology Assessment Division (MD-67), Research Triangle Park, NC 27711.
R M
RM
Zucker
K M
KM
Daniel
E J
EJ
Massaro
S J
SJ
Karafas
L L
LL
Degn
W K
WK
Boyes
D002460:Cell Line; D005434:Flow Cytometry / Q000379:methods*; D005453:Fluorescence; D006801:Humans; D053768:Metal Nanoparticles / Q000032:analysis*; D012834:Silver; D020349:Surface Plasmon Resonance
D012834:Silver
dark‐field microscopy; far red fluorescence; flow cytometry; light scatter; plasmonic surface fluorescence; silver; silver nanoparticles; surface enhanced fluorescence; titanium dioxide