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A Nanoparticle-based Sensor Platform for Cell Tracking and Status/Function Assessment.

Yeo D, Wiraja C, Chuah YJ, Gao Y, Xu C - Sci Rep (2015)

Bottom Line: Upon intracellular entry, nanosensors reside within the cell cytoplasm, serving as a depot to continuously release sensor molecules for up to 30 days.When the biomarker(s) is expressed, a detectable signal is generated (On).As a proof-of-concept, three nanosensor formulations were synthesized to monitor cell viability, secretion of nitric oxide, and β-actin mRNA expression.

View Article: PubMed Central - PubMed

Affiliation: School of Chemical &Biomedical Engineering, Nanyang Technological University, Singapore.

ABSTRACT
Nanoparticles are increasingly popular choices for labeling and tracking cells in biomedical applications such as cell therapy. However, all current types of nanoparticles fail to provide real-time, noninvasive monitoring of cell status and functions while often generating false positive signals. Herein, a nanosensor platform to track the real-time expression of specific biomarkers that correlate with cell status and functions is reported. Nanosensors are synthesized by encapsulating various sensor molecules within biodegradable polymeric nanoparticles. Upon intracellular entry, nanosensors reside within the cell cytoplasm, serving as a depot to continuously release sensor molecules for up to 30 days. In the absence of the target biomarkers, the released sensor molecules remain 'Off'. When the biomarker(s) is expressed, a detectable signal is generated (On). As a proof-of-concept, three nanosensor formulations were synthesized to monitor cell viability, secretion of nitric oxide, and β-actin mRNA expression.

No MeSH data available.


Related in: MedlinePlus

Viability nanosensors for labeling mesenchymal stem cells (MSCs).(A) Cumulative release of CAM from nanosensors over 28 days in PBS at 37 °C. (B) Fluorescence signal of supernatants in A before and after the esterase treatment. (C) Flow cytometry analysis of MSCs before (grey) and after (red) the labeling with 3 mg/ml viability nanosensor. (D) Nanosensor labeled MSCs stained with Hoechst 33342 (from left to right: nanosensors, nuclei, merged image). Scale bars are 100 μm. (E) Confocal images of nanosensor labeled MSCs stained with DiI and Hoechst 33342 (from left to right: nanosensors, plasma membrane, nuclei, merged image); Z-projection (main), YZ-plane (right), XZ-plane (below). Scale bars are 20 μm. Values are mean ± SD, N = 4.
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f2: Viability nanosensors for labeling mesenchymal stem cells (MSCs).(A) Cumulative release of CAM from nanosensors over 28 days in PBS at 37 °C. (B) Fluorescence signal of supernatants in A before and after the esterase treatment. (C) Flow cytometry analysis of MSCs before (grey) and after (red) the labeling with 3 mg/ml viability nanosensor. (D) Nanosensor labeled MSCs stained with Hoechst 33342 (from left to right: nanosensors, nuclei, merged image). Scale bars are 100 μm. (E) Confocal images of nanosensor labeled MSCs stained with DiI and Hoechst 33342 (from left to right: nanosensors, plasma membrane, nuclei, merged image); Z-projection (main), YZ-plane (right), XZ-plane (below). Scale bars are 20 μm. Values are mean ± SD, N = 4.

Mentions: To examine the feasibility of this idea, nanosensors designed to assess cell viability (i.e. viability nanosensors) were first synthesized by encapsulating CAM within poly(lactic-co-glycolic acid) (PLGA) NPs in the size range of 500 nm–1 μm (Supplementary Fig. S1a,b). When placed in phosphate buffered saline (PBS), a 10% release was observed during day 1 and 90% of encapsulated CAM molecules was subsequently released over 28 days (Fig. 2a). This was quantified using an absorption-concentration calibration curve of free CAM (Supplementary Fig. S1c). CAM is known to be weakly fluorescent, but converts to strongly fluorescent calcein in the presence of esterases. When the aqueous supernatant was treated with esterases, we observed a ~20% increase of fluorescent intensity (Fig. 2b, Supplementary Fig. S1d). Compared to freshly constituted CAM (over ~120% enhancement with the esterase treatment, Supplementary Fig. S1e,f), the released CAM sensors exhibited reduced signal enhancement. This reduced functionality was in all likelihood, a result of spontaneous CAM hydrolysis in aqueous solution12.


A Nanoparticle-based Sensor Platform for Cell Tracking and Status/Function Assessment.

Yeo D, Wiraja C, Chuah YJ, Gao Y, Xu C - Sci Rep (2015)

Viability nanosensors for labeling mesenchymal stem cells (MSCs).(A) Cumulative release of CAM from nanosensors over 28 days in PBS at 37 °C. (B) Fluorescence signal of supernatants in A before and after the esterase treatment. (C) Flow cytometry analysis of MSCs before (grey) and after (red) the labeling with 3 mg/ml viability nanosensor. (D) Nanosensor labeled MSCs stained with Hoechst 33342 (from left to right: nanosensors, nuclei, merged image). Scale bars are 100 μm. (E) Confocal images of nanosensor labeled MSCs stained with DiI and Hoechst 33342 (from left to right: nanosensors, plasma membrane, nuclei, merged image); Z-projection (main), YZ-plane (right), XZ-plane (below). Scale bars are 20 μm. Values are mean ± SD, N = 4.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4593999&req=5

f2: Viability nanosensors for labeling mesenchymal stem cells (MSCs).(A) Cumulative release of CAM from nanosensors over 28 days in PBS at 37 °C. (B) Fluorescence signal of supernatants in A before and after the esterase treatment. (C) Flow cytometry analysis of MSCs before (grey) and after (red) the labeling with 3 mg/ml viability nanosensor. (D) Nanosensor labeled MSCs stained with Hoechst 33342 (from left to right: nanosensors, nuclei, merged image). Scale bars are 100 μm. (E) Confocal images of nanosensor labeled MSCs stained with DiI and Hoechst 33342 (from left to right: nanosensors, plasma membrane, nuclei, merged image); Z-projection (main), YZ-plane (right), XZ-plane (below). Scale bars are 20 μm. Values are mean ± SD, N = 4.
Mentions: To examine the feasibility of this idea, nanosensors designed to assess cell viability (i.e. viability nanosensors) were first synthesized by encapsulating CAM within poly(lactic-co-glycolic acid) (PLGA) NPs in the size range of 500 nm–1 μm (Supplementary Fig. S1a,b). When placed in phosphate buffered saline (PBS), a 10% release was observed during day 1 and 90% of encapsulated CAM molecules was subsequently released over 28 days (Fig. 2a). This was quantified using an absorption-concentration calibration curve of free CAM (Supplementary Fig. S1c). CAM is known to be weakly fluorescent, but converts to strongly fluorescent calcein in the presence of esterases. When the aqueous supernatant was treated with esterases, we observed a ~20% increase of fluorescent intensity (Fig. 2b, Supplementary Fig. S1d). Compared to freshly constituted CAM (over ~120% enhancement with the esterase treatment, Supplementary Fig. S1e,f), the released CAM sensors exhibited reduced signal enhancement. This reduced functionality was in all likelihood, a result of spontaneous CAM hydrolysis in aqueous solution12.

Bottom Line: Upon intracellular entry, nanosensors reside within the cell cytoplasm, serving as a depot to continuously release sensor molecules for up to 30 days.When the biomarker(s) is expressed, a detectable signal is generated (On).As a proof-of-concept, three nanosensor formulations were synthesized to monitor cell viability, secretion of nitric oxide, and β-actin mRNA expression.

View Article: PubMed Central - PubMed

Affiliation: School of Chemical &Biomedical Engineering, Nanyang Technological University, Singapore.

ABSTRACT
Nanoparticles are increasingly popular choices for labeling and tracking cells in biomedical applications such as cell therapy. However, all current types of nanoparticles fail to provide real-time, noninvasive monitoring of cell status and functions while often generating false positive signals. Herein, a nanosensor platform to track the real-time expression of specific biomarkers that correlate with cell status and functions is reported. Nanosensors are synthesized by encapsulating various sensor molecules within biodegradable polymeric nanoparticles. Upon intracellular entry, nanosensors reside within the cell cytoplasm, serving as a depot to continuously release sensor molecules for up to 30 days. In the absence of the target biomarkers, the released sensor molecules remain 'Off'. When the biomarker(s) is expressed, a detectable signal is generated (On). As a proof-of-concept, three nanosensor formulations were synthesized to monitor cell viability, secretion of nitric oxide, and β-actin mRNA expression.

No MeSH data available.


Related in: MedlinePlus