Detection of silver nanoparticles in cells by flow cytometry using light scatter and far‐red fluorescence

Abstract

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.

Semi‐quantitative estimation of cellular SiO2nanoparticles using flow cytometry combined with X‐ray fluorescence measurements

“…Another study also reported that the SSC intensity of FCM measurements is closely related to the internalization of Fe 3 O 4 , TiO 2 , and Ag NPs (14,16,17). Particularly, Zucker et al (16,17) have confirmed that the cellular associations of TiO 2 and Ag NPs are closely related to the SSC signals of FCM by correlating FCM results with images from dark field microscopy. Due to this unique feature, FCM SSC measurements for the analysis of cellular NP association can be used as a simple, sensitive, and cost-effective technique for initial screening of the NP's potential to interact with biological cells and tissues (15)(16)(17)(18)(19).…”

“…Particularly, Zucker et al (16,17) have confirmed that the cellular associations of TiO 2 and Ag NPs are closely related to the SSC signals of FCM by correlating FCM results with images from dark field microscopy. Due to this unique feature, FCM SSC measurements for the analysis of cellular NP association can be used as a simple, sensitive, and cost-effective technique for initial screening of the NP's potential to interact with biological cells and tissues (15)(16)(17)(18)(19). However, although, this FCM measurement of cellular NPs could be a general approach useful for detecting cell-associated NPs, this FCM approach has only been used to qualitatively observe NP association with mammalian cells rather than quantitative measurement of cellular NP contents.…”

“…Therefore, these light scattering data from the FCM measurements can provide useful insight into the cellular association with NPs. For example, it was recently reported that the mean intensity of SSC is proportional to the concentration of NPs inside the cells (15)(16)(17)(18). Another study also reported that the SSC intensity of FCM measurements is closely related to the internalization of Fe 3 O 4 , TiO 2 , and Ag NPs (14,16,17).…”

“…For example, it was recently reported that the mean intensity of SSC is proportional to the concentration of NPs inside the cells (15)(16)(17)(18). Another study also reported that the SSC intensity of FCM measurements is closely related to the internalization of Fe 3 O 4 , TiO 2 , and Ag NPs (14,16,17). Particularly, Zucker et al (16,17) have confirmed that the cellular associations of TiO 2 and Ag NPs are closely related to the SSC signals of FCM by correlating FCM results with images from dark field microscopy.…”

High throughput toxicity screening and intracellular detection of nanomaterials

“…It was demonstrated that TiO 2 78, 79, 80 and Ag81 NMs induce an increase of SS signal in cell populations after exposure to both NMs. SS indicates the granularity of the cells and is strongly linked to their content of NMs.…”