TITLE: Synthesis and Characterization of Silver Nanoparticles using Sodium Borohydride as a Reducing Agent  
AUTHORS: Sean  Byrne McDonnell, Sebnem  Gunes, Alan  Casey, James Curtin, Eline  Manaloto

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Silver nanoparticles (AgNP) have a multitude of uses, which range from anti-microbial to anti-cancerous etc. They are sometimes preferred to other metal-based nanoparticles due to their attractive physicochemical properties.  This protocol is a standardized method of producing and characterising PVA coated AgNPs which range from 10 to 20 nm in diameter. These particles are suited to lab based in-vivo and in-vitro assays. The addition of PVA to AgNPs is to increase the stability of the nanoparticles. </div></div>

[STEPS]
?. [Synthesis]
Add  of   to an Erlenmeyer flask.
30 mL
?. [Synthesis]
Insert the Erlenmeyer flask containing  of sodium borohydride into the previously set up ice bath containing the stir plate and allow to cool for
30 mL
?. [Synthesis]
Place a magnetic mixing rod into the Erlenmeyer flask and start the stir plate to initiate mixing.
?. [Synthesis]
In another Erlenmeyer flask, transfer   of the freshly prepared  along with  of  a 1% PVA solution.
2 mL
2 mL
1% Polyvinyl Alcohol in Millipore water (Simplicity 185, 18.2 MΩ.cm at 25°C resistivity)
?. [Synthesis]
At a rate of one drop per second, drip  of the AgNO3/PVA solution into the chilled  solution, maintaining the cooling and mixing simultaneously.
2 mL
?. [Synthesis]
When the AgNO3/PVA solution has been added to the  solution, stop mixing immediately.
Once the reaction has been stopped, note that a yellow solution should be formed. This indicates that PVA-AgNP formation has occurred.
?. [Purification ]
Transfer  of the PVA-AgNP solution, along with the excess into an ultrafiltration tube  of 3kDa.
20 mL
3kDa Ultrafiltration tube used in this protocol was from Sartorius. This step may need to be repeated for volumes >20mL.
?. [Purification ]
Transfer the tube into a centrifuge and run at
Centrifuge: 5000 33, 40 min
?. [Purification ]
Post centrifugation, smaller NPs  (>5μm in diameter) and excess water (including unwanted reactants) can be discarded from the ultrafiltration tube.
The smaller AgNPs will be seen at the clustered at the bottom of the tube.
?. [Purification ]
The PVA-AgNP concentrate will display a dark brown colour, this is then collected using a dropper.
?. [Purification ]
The harvested AgNPs must then be stored at fridge temperature (Approx. 4°C)  away from direct sunlight.
?. [Characterisation]
AgNP samples are prepared using a  1 in 10 dilution by adding 1 ml concentrated PVA-AgNP to 9 mL of millipore water
?. [Characterisation]
For primary characterisation,  of PVA-AgNP's are placed into a cuvette, a UV-Vis spectrum is ran from 300nm-600nm.
3.5 mL
Strong absorbance at 400nm indicates 10mm AgNP synthesis.
?. [Characterisation]
Using a DLS instrument (Malvern Instruments, UK) particle size range is measured by placing   PVA-AgNP soloution into a 12mm o.d. square polystyrene cuvette and ran as per manufactuer specification.
3.5 mL
DLS Measures hydrodynamic size, including both the PVA coating the AgNP's.Zetasizer software was used to calculate the size distribution.
?. [Characterisation]
The DLS instrument was again used to measure the Zeta Potential of the AgNPs, this was performed on the same sample specifications for the previous DLS sample  .
Zeta potential measures the potential for the particles to flocculate, with a high negative or positive charge causing them to repel one another.
?. [Characterisation]
STEM Characterization,  of PVA-AgNP aqueous solution is placed onto a carbon-coated copper plate and allowed to dry. Particles are then analyzed visually for dispersion and shape.
3 µl
Drying allows for the highest yield of particles on the grid.
?. [Characterisation]
Using the previously prepared calibration solutions (1-5 ug AgNP/mL) the concentration of the prepared AgNP solution was calculated using AAS (Varian SpectrAA 200).