Nanosensors were synthesized through single or double emulsion methods4 (link). For viability and NO nanosensors, 250 μg CAM or 1 mg DAF-FM DA in 2 ml chloroform was mixed with 100 mg PLGA (50:50) at 4 °C. This mixture was added dropwise to 3% polyvinyl alcohol (PVA) aqueous solution and homogenized (Tissue Master 125, Omni International) for 60 s. The emulsion was placed in the chemical hood to evaporate chloroform for 3 hrs. Finally, nanosensors were collected through centrifugation (6,000 rpm) and washed 3× with double-distilled water before freezing-drying.
To generate β-actin mRNA nanosensors, 0.1 nmol MBs were first dissolved in 100 μl double-distilled water and homogenized into 500 μl chloroform containing 10 mg of 50:50 PLGA with 1% Span® 80. This water-oil emulsion was further homogenized with 3% PVA to form a water-oil-water double emulsion. All other fabrication steps were similar to the single emulsion as above.
To generate β-actin mRNA nanosensors, 0.1 nmol MBs were first dissolved in 100 μl double-distilled water and homogenized into 500 μl chloroform containing 10 mg of 50:50 PLGA with 1% Span® 80. This water-oil emulsion was further homogenized with 3% PVA to form a water-oil-water double emulsion. All other fabrication steps were similar to the single emulsion as above.
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