This work was supported by Agency for Defense Development (Grant No. UD170039ID).

1. Preparation of Poly(PFPA) Homopolymer
<ol>
	<li>Recrystallization of AIBN
	<ol>
		<li>Combine 5 g of 2,2&#8217;-azobis(2-methylpropionitrile) (AIBN) with 25 mL of methanol in a 250 mL beaker. Immerse the beaker in a 60 &#176;C oil bath, then vigorously stir the mixture with a stir bar until AIBN is fully dissolved.</li>
		<li>Filter the warm solution through filter paper (5-8 &#956;m particle retention) and store the filtrate at 4 &#176;C to allow the crystals to form slowly.</li>
		<li>Collect the recrystallized AIBN by filtration. Combine the collected product with 25 mL of fresh methanol and repeat the recrystallization process.</li>
		<li>Dry the 2x recrystallized AIBN in a vacuum oven at room temperature (RT) overnight. Store the product in the dark at &#60; -10 &#176;C.</li>
	</ol>
	</li>
	<li>Synthesis of benzyl dithiobenzoate25
	<ol>
		<li>Prepare a 500 mL three-neck round-bottom flask equipped with a magnetic stir bar, a refluxing condenser, a dropping funnel, and a rubber septum. Connect the flask to the nitrogen gas line through the refluxing condenser and flush out the inside air with nitrogen. Insert a thermometer through the septum. Add 41 mL (0.041 mol) of 1 M solution of phenylmagnesium bromide in tetrahydrofuran (THF) via a syringe through the same septum.</li>
		<li>Warm the phenylmagnesium bromide solution to 40 &#176;C in an oil bath. Then add 3.1 g (0.041 mol) of carbon disulfide through the dropping funnel slowly, maintaining the solution temperature at 40 &#176;C.</li>
		<li>Add 7.1 g (0.042 mol) of benzyl bromide to the resultant mixture through the dropping funnel over 15 min. Increase the reaction temperature to 50 &#176;C. Continue stirring at this temperature for 45 min.</li>
		<li>Transfer the reaction mixture into a separatory funnel and dilute with 15 mL of ice cold water. Extract the product by adding 15 mL of diethyl ether and remove the lower water layer. Repeat the extraction with diethyl ether two more times.</li>
		<li>Wash the combined organic phases with copious amount of water, then brine (solution of 50% (w/v) NaCl in water) and dry the product over anhydrous magnesium sulfate.</li>
		<li>Remove the solvent in vacuum at 35&#160;&#176;C using a rotary evaporator.</li>
		<li>Purify the product by column chromatography using 400 mL of silica gel (pore size 60 &#197;, 63-200 mesh particle size) and petroleum ether as the eluent, yielding 5 g of benzyl dithiobenzoate (BDB) as red oil. Confirm the product purity by 1H NMR (400 MHz, CDCl3): &#948;&#160;8.02-7.99 (2H, m), 7.55-7.50 (1H, m), 7.41-7.29 (7H, m), 4.60 (2H, s).</li>
	</ol>
	</li>
	<li>Synthesis of poly(PFPA) via RAFT polymerization9,26
	<ol>
		<li>Commercially available PFPA monomer contains small amount of inhibitors. Before polymerization, remove the inhibitors by passing the monomer through a disposable syringe packed with basic alumina.</li>
		<li>Add 0.4 mg (0.0024 mmol) of recrystallized AIBN, 4.3 mg (0.018 mmol) of BDB, 1012 mg (4.25 mmol) of inhibitor-free PFPA, and 0.7 mL of anhydrous anisole to a 20 mL Schlenk flask.</li>
		<li>Connect the flask to Schlenk line and degas the mixture with at least three freeze-pump-thaw cycles. Briefly, freeze the reaction mixture in a liquid nitrogen bath. Apply vacuum to remove the gas in headspace. Seal the flask then remove away from liquid nitrogen to allow the content to thaw at RT.</li>
		<li>Place the flask in a 70 &#176;C oil bath, and react for 4 h under N2 purge.</li>
		<li>To terminate the reaction, remove the flask from the oil bath and expose the reaction content to air.</li>
		<li>Precipitate the polymer in cold methanol, then dry the recovered polymer in a vacuum oven at 40 &#176;C overnight.</li>
		<li>To measure the polymer molecular weight, use gel permeation chromatography (GPC). Use THF as the mobile phase at 35 &#176;C with a 1 mL/min flow rate and construct the calibration curve using monodisperse polystyrene standards. To acquire GPC measurement, dissolve the polymer in THF (1-2 mg/mL) and filter through 0.2 &#956;m disposable polytetrafluoroethylene (PTFE) filter. Inject 100 &#956;L of the sample into the GPC instrument. Convert the measured sample retention time to molecular weight using the polystyrene calibration curve.</li>
	</ol>
	</li>
</ol>
2. Preparation of Poly(PFPA) Functionalized SiO2 Beads
<ol>
	<li>Treatment of SiO2 beads with APTMS
	<ol>
		<li>SiO2 particles are available in the form of a 5% (w/v) aqueous suspension. Combine 0.8 mL of SiO2 suspension with 40 mg of APTMS and 8 mL of methanol in a 20 mL scintillation vial equipped with a stir bar.</li>
		<li>Allow the reaction to proceed at RT for 5 h with vigorous stirring.</li>
		<li>Transfer the solution to a conical tube. To isolate the APTMS functionalized SiO2 beads, centrifuge the solution at 10,000 x g for 5 min, then remove the supernatant. Wash the beads by re-dispersing them in 3 mL of fresh methanol. Shake the tube by hand for mixing, but if necessary, improve the dispersion by sonication in a water bath for a few seconds. Centrifuge the beads at 10,000 x g for 5 min. Remove the supernatant and repeat the wash step one more time.</li>
		<li>Combine the methanol washed SiO2 beads with 3 mL of dimethyl sulfoxide (DMSO). Shake the mixture by hand, or if necessary sonicate for a few seconds, until the beads are fully dispersed in DMSO. Centrifuge the beads at 10,000 x g for 5 min, then remove the supernatant. Repeat the step to ensure complete solvent exchange from methanol to DMSO. 
		NOTE: The final suspension contains the APTMS functionalized SiO2 beads dispersed in 4 mL of DMSO.</li>
		<li>To check the particle size distribution, perform DLS analysis. Take one drop of the suspension prepared in Step 2.1.4 and place into a disposable UV cuvette. Dilute the sample by filling the cuvette with fresh DMSO until it is 2/3 full. Insert the sample into the cell holder to begin data acquisition. For particle size measurement, use the following setup parameters: Temperature: 25 &#176;C; Equilibration Time: 120 s; Measurement Duration: Automatic.</li>
		<li>To check the surface composition, perform XPS analysis. Dry a small sample from the suspension prepared in Step 2.1.4 in vacuum oven at 40 &#176;C overnight. Take the dried polymer and pack evenly onto a 0.5 cm x 0.5 cm sample holder. Load the sample into the high vacuum chamber (10-8 torr) and begin data acquisition. For the particular XPS instrument used, generate the photoelectrons using a monochromatic Al K&#945;&#160;X-ray operated at 15 kV and 6.7 mA, and collect using hybrid mode magnification with the analyzer at a 50 eV pass energy for high resolution spectra, and a 100 eV pass energy for elemental surveys.</li>
	</ol>
	</li>
	<li>Grafting poly(PFPA) to APTMS functionalized SiO2 beads
	<ol>
		<li>Prepare the poly(PFPA) solution by dissolving 20 mg of poly(PFPA) in 2 mL of DMSO in a 20 mL scintillation vial. 
		NOTE: In this study, a relatively low molecular weight poly(PFPA) (20 kg/mol) is used. Thus, despite the high polymer concentration (10 mg/mL), no evidence of polymer crosslinking is observed. If a higher molecular weight polymer is used, then polymer solution concentration may need to be adjusted to avoid possible crosslinking.</li>
		<li>Add 1 mL of APTMS functionalized SiO2 beads suspended in DMSO (from Step 2.1.4) to the poly(PFPA) solution. React at RT for 1 h with vigorous stirring.</li>
		<li>Isolate the poly(PFPA) grafted SiO2 beads by centrifugation at 10,000 x g for 5 min, followed by the removal of the supernatant. Wash the beads by adding 3 mL of DMSO and mix by either shaking with hand or few seconds of sonication. Centrifuge the beads at 10,000 x g for 5 min, then remove the supernatant. Repeat washing of the poly(PFPA) grafted SiO2 beads with DMSO twice.</li>
		<li>Wash the beads two times more with triple distilled water (TDW). In this step, combine the beads with 3 mL of TDW, then mix by shaking with hand or few seconds of sonication. Centrifuge the beads at 10,000 x g for 5 min, then remove the supernatant.</li>
		<li>To check the particle size distribution, perform DLS following the procedure described in Step 2.1.5. To check the surface chemistry, perform XPS following the procedure described in Step 2.1.6.</li>
	</ol>
	</li>
</ol>
3. Preparation of SiO2 Beads Grafted with PEG-Substituted Poly(PFPA)
<ol>
	<li>To prepare the poly(PFPA) solution, dissolve 20 mg of poly(PFPA) in 2 mL of DMSO in a 20 mL scintillation vial.</li>
	<li>To prepare PEG solution, dissolve amine-functionalized PEG in 1 mL of DMSO. The exact amount of PEG used is determined by the desired degree of PFP substitution, determined by the equation shown below: 
	Amount of amino-PEG (g/g-poly(PFPA)) = (N_poly(PFPA) x % PEG-Sub) x (MW_PEG / MW_poly(PFPA)) 
	where N_poly(PFPA) = poly(PFPA) degree of polymerization 
	% PEG-Sub = percent PEG substitution 
	MW_PEG = molecular weight of amino-PEG 
	MW_ poly(PFPA) = molecular weight of poly(PFPA)</li>
	<li>Transfer the PEG solution to the poly(PFPA) solution. React at RT for 1 h with vigorous stirring.</li>
	<li>To prepare APTMS functionalized SiO2 beads suspended in DMSO, follow the same steps shown in Step 2.1. Transfer 1 mL of the bead suspension into the PEG-substituted poly(PFPA) solution prepared in Step 3.3. Allow the grafting between poly(PFPA) and APTMS functionalized SiO2 beads to proceed at RT for 1 h with vigorous stirring.</li>
	<li>Isolate the beads by centrifugation at 10,000 x g for 5 min, followed by the removal of the supernatant. Wash the beads by adding 3 mL of DMSO and mix by either shaking with hand or few seconds of sonication. Centrifuge the beads at 10,000 x g for 5 min, then remove the supernatant. Repeat the DMSO wash twice.</li>
	<li>Wash the beads two times more with TDW. In this step, combine the beads with 3 mL TDW, then mix by shaking with hand or few seconds of sonication. Centrifuge the beads at 10,000 x g for 5 min, then remove the supernatant.</li>
	<li>Dry the beads at 40 &#176;C in a vacuum oven overnight.</li>
</ol>
4. Antibody Immobilization on Poly(PFPA) Grafted SiO2 Beads
NOTE: The same procedure is used regardless of percent PEG substitution on poly(PFPA). Prepare phosphate buffered saline (PBS) by dissolving PBS tablet in TDW. Prepare 0.1% (v/v) phosphate buffered saline with Tween-20 (PBST) by adding 1/1000 of Tween-20 to PBS.
<ol>
	<li>Add 5 mg of poly(PFPA) grafted SiO2 beads to a 1.5 mL microcentrifuge tube.</li>
	<li>Wash the beads by adding 800 &#181;L of PBS and mix well by vortexing. Centrifuge the beads at 10,000 x g at RT for 1 min. Remove the supernatant and repeat the wash step three times.</li>
	<li>Add 350 &#181;L of fresh PBS, 50 &#181;L 0.1% (v/v) PBST, and 6.67 &#181;g of the antibody. Incubate ~20 h on a rotator at 4 &#176;C.</li>
	<li>Wash the beads to remove unbound antibodies. Centrifuge the beads at 400 x g and 4 &#176;C for 1 min. Remove the supernatant and add 400 &#181;L of lysis buffer carefully. Gently re-suspend the beads by pipetting up and down for five times. 
	NOTE: Lysis buffer used to wash the beads should be the same one used during cell lysis and IP, except that the addition of dithiothreitol and protease inhibitor are optional, (see Step 5).</li>
	<li>Repeat this wash step three times. After the final wash, remove the supernatant as much as possible.</li>
</ol>
5. Cell Lysis and Immunoprecipitation
<ol>
	<li>Preparation of lysis buffer and wash buffer
	<ol>
		<li>Prepare the lysis buffer (50 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.5% (v/v) NP-40, 10% (v/v) glycerol, 1 mM dithiothreitol (DTT), and protease inhibitor cocktail).</li>
		<li>Prepare the wash buffer (50 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 % (v/v) NP-40, and 10% (v/v) glycerol).</li>
		<li>Store the buffer solutions at 4 &#176;C.</li>
	</ol>
	</li>
	<li>Preparation of the cells
	<ol>
		<li>Seed the cells (HeLa cells) one or two days before IP experiment, and grow the cells at 37 &#176;C and 5% CO2.</li>
		<li>Collect about 1.4 x 107 cells with a cell scraper and transfer into a 15 mL conical tube. Centrifuge the cells at 380 x g at RT for 3 min. Remove the supernatant and re-suspend with 1 mL of cold PBS and transfer into a 1.5 mL microcentrifuge tube.</li>
		<li>Centrifuge the cells at 10,000 x g at 4 &#176;C for 30 s. Remove the supernatant cleanly. Cell pellets can be stored at -80 &#176;C after removing the supernatant.</li>
	</ol>
	</li>
	<li>Preparation of cell lysates
	<ol>
		<li>Re-suspend the cell pellet with 400 &#181;L of the lysis buffer. Sonicate the cells using an ultrasonicator.</li>
		<li>After the sonication, vortex briefly and centrifuge the lysate at 20,000 x g at 4 &#176;C for 10 min.</li>
		<li>Transfer the supernatant to a new 1.5 mL centrifuge tube.</li>
	</ol>
	</li>
	<li>Immunoprecipitation
	<ol>
		<li>Transfer 300 &#181;L of cell lysate to previously prepared antibody incubated poly(PFPA) grafted SiO2 beads. Retain 30 &#181;L of the cell lysate as the input sample in a new microcentrifuge tube. Store the input sample at 4 &#176;C. 
		NOTE: The total amount of protein in cell lysate should be approximately 4 mg.</li>
		<li>Incubate the lysate/beads mixture for 3 h on a rotator at 4 &#176;C.</li>
		<li>Centrifuge the mixture at 400 x g at 4 &#176;C for 1 min. Remove the supernatant and add 400 &#181;L of wash buffer carefully. Gently re-suspend the beads by pipetting up and down about five times.</li>
		<li>Repeat this wash step three times. After the final wash, remove the supernatant as much as possible.</li>
		<li>Prepare 2x sodium dodecyl sulfate (SDS) loading dye (25% (v/v) glycerol, 0.1% (w/v) bromo phenol blue (BPB), 60 mM Tris-HCl (pH 6.8), 2% (w/v) SDS, and 2.75 mM 2-mercaptoethanol). Store 2x SDS loading dye at -20 &#176;C. Add 30 &#181;L of 2x SDS loading dye to the beads and the stored input sample, and heat them for 10 min at 95 &#176;C.</li>
		<li>After heating, analyze the sample using Western blotting27, or store the sample at -20 &#176;C.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Johnsson, B., L&#246;f&#229;s, S., Lindquist, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immobilization+of+proteins+to+a+carboxymethyldextran-modified+gold+surface+for+biospecific+interaction+analysis+in+surface+plasmon+resonance+sensors.">Immobilization of proteins to a carboxymethyldextran-modified gold surface for biospecific interaction analysis in surface plasmon resonance sensors.</a> Analytical Biochemistry. 198 (2), 268-277 (1991).</li><li>Kurzawa, C., Hengstenberg, A., Schuhmann, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immobilization+method+for+the+preparation+of+biosensors+based+on+pH+shift-induced+deposition+of+biomolecule-containing+polymer+films.">Immobilization method for the preparation of biosensors based on pH shift-induced deposition of biomolecule-containing polymer films.</a> Analytical Chemistry. 74 (2), 355-361 (2002).</li><li>You, C. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+and+identification+of+proteins+using+nanoparticle-fluorescent+polymer+'chemical+nose'+sensors.">Detection and identification of proteins using nanoparticle-fluorescent polymer 'chemical nose' sensors.</a> Nature Nanotechnology. 2 (5), 318-323 (2007).</li><li>Roberts, M. W., Ongkudon, C. M., Forde, G. M., Danquah, M. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Versatility+of+polymethacrylate+monoliths+for+chromatographic+purification+of+biomolecules.">Versatility of polymethacrylate monoliths for chromatographic purification of biomolecules.</a> Journal of Separation Science. 32 (15-16), 2485-2494 (2009).</li><li>Sandison, M. E., Cumming, S. A., Kolch, W., Pitt, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On-chip+immunoprecipitation+for+protein+purification.">On-chip immunoprecipitation for protein purification.</a> Lab on a Chip. 10 (20), 2805-2813 (2010).</li><li>Das, A., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activated+Ester+Containing+Polymers:+Opportunities+and+Challenges+for+the+Design+of+Functional+Macromolecules.">Activated Ester Containing Polymers: Opportunities and Challenges for the Design of Functional Macromolecules.</a> Chemical Reviews. 116 (3), 1434-1495 (2016).</li><li>Choi, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functionalization+and+patterning+of+reactive+polymer+brushes+based+on+surface+reversible+addition+and+fragmentation+chain+transfer+polymerization.">Functionalization and patterning of reactive polymer brushes based on surface reversible addition and fragmentation chain transfer polymerization.</a> Journal of Polymer Science Part A: Polymer Chemistry. 50 (19), 4010-4018 (2012).</li><li>Kessler, D., Jochum, F. D., Choi, J., Char, K., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reactive+surface+coatings+based+on+polysilsesquioxanes:+universal+method+toward+light-responsive+surfaces.">Reactive surface coatings based on polysilsesquioxanes: universal method toward light-responsive surfaces.</a> ACS Applied Materials &amp; Interfaces. 3 (2), 124-128 (2011).</li><li>Son, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Penetration+and+exchange+kinetics+of+primary+alkyl+amines+applied+to+reactive+poly(pentafluorophenyl+acrylate)+thin+films.">Penetration and exchange kinetics of primary alkyl amines applied to reactive poly(pentafluorophenyl acrylate) thin films.</a> Polymer Journal. 48 (4), 487-495 (2016).</li><li>Kessler, D., Roth, P. J., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reactive+surface+coatings+based+on+polysilsesquioxanes:+controlled+functionalization+for+specific+protein+immobilization.">Reactive surface coatings based on polysilsesquioxanes: controlled functionalization for specific protein immobilization.</a> Langmuir. 25 (17), 10068-10076 (2009).</li><li>Kessler, D., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reactive+surface+coatings+based+on+polysilsesquioxanes:+defined+adjustment+of+surface+wettability.">Reactive surface coatings based on polysilsesquioxanes: defined adjustment of surface wettability.</a> Langmuir. 25 (24), 14200-14206 (2009).</li><li>Kessler, D., Nilles, K., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modular+approach+towards+multi-functional+surfaces+with+adjustable+and+dual-responsive+wettability+using+a+hybrid+polymer+toolbox.">Modular approach towards multi-functional surfaces with adjustable and dual-responsive wettability using a hybrid polymer toolbox.</a> Journal of Materials Chemistry. 19 (43), 8184-8189 (2009).</li><li>Eberhardt, M., Mruk, R., Zentel, R., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+of+pentafluorophenyl(meth)acrylate+polymers:+new+precursor+polymers+for+the+synthesis+of+multifunctional+materials.">Synthesis of pentafluorophenyl(meth)acrylate polymers: new precursor polymers for the synthesis of multifunctional materials.</a> European Polymer Journal. 41 (7), 1569-1575 (2005).</li><li>Jochum, F. D., Forst, F. R., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PNIPAM+copolymers+containing+light-responsive+chromophores:+a+method+toward+molecular+logic+gates.">PNIPAM copolymers containing light-responsive chromophores: a method toward molecular logic gates.</a> Macromolecular Rapid Communications. 31 (16), 1456-1461 (2010).</li><li>Schattling, P., Pollmann, I., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+of+CO2-responsive+polymers+by+post-polymerization+modification.">Synthesis of CO2-responsive polymers by post-polymerization modification.</a> Reactive &amp; Functional Polymers. 75, 16-21 (2014).</li><li>He, L., Szameit, K., Zhao, H., Hahn, U., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postpolymerization+modification+using+less+cytotoxic+activated+ester+polymers+for+the+synthesis+of+biological+active+polymers.">Postpolymerization modification using less cytotoxic activated ester polymers for the synthesis of biological active polymers.</a> Biomacromolecules. 15 (8), 3197-3205 (2014).</li><li>Arnold, R. M., McNitt, C. D., Popik, V. V., Locklin, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+grafting+of+poly(pentafluorophenyl+acrylate)+onto+oxides:+versatile+substrates+for+reactive+microcapillary+printing+and+self-sorting+modification.">Direct grafting of poly(pentafluorophenyl acrylate) onto oxides: versatile substrates for reactive microcapillary printing and self-sorting modification.</a> Chemical Communications. 50 (40), 5307-5309 (2014).</li><li>Son, H., Ku, J., Kim, Y., Li, S., Char, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Amine-Reactive+Poly(pentafluorophenyl+acrylate)+Brush+Platforms+for+Cleaner+Protein+Purification.">Amine-Reactive Poly(pentafluorophenyl acrylate) Brush Platforms for Cleaner Protein Purification.</a> Biomacromolecules. 19 (3), 951-961 (2018).</li><li>Cullen, S. E., Schwartz, B. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+method+for+isolation+of+H-2+and+Ia+alloantigens+with+immunoprecipitation+induced+by+protein+A-bearing+staphylococci.">An improved method for isolation of H-2 and Ia alloantigens with immunoprecipitation induced by protein A-bearing staphylococci.</a> The Journal of Immunology. 117 (1), 136-142 (1976).</li><li>Sisson, T. H., Castor, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+method+for+immobilizing+IgG+antibodies+on+protein+A-agarose.">An improved method for immobilizing IgG antibodies on protein A-agarose.</a> Journal of Immunology Methods. 127 (2), 215-220 (1990).</li><li>Peritz, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunoprecipitation+of+mRNA-protein+complexes.">Immunoprecipitation of mRNA-protein complexes.</a> Nature Protocols. 1 (2), 577-580 (2006).</li><li>Zhang, Z., Chen, S., Jiang, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dual-functional+biomimetic+materials:+nonfouling+poly+(carboxybetaine)+with+active+functional+groups+for+protein+immobilization.">Dual-functional biomimetic materials: nonfouling poly (carboxybetaine) with active functional groups for protein immobilization.</a> Biomacromolecules. 7 (12), 3311-3315 (2006).</li><li>Yao, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NHS-ester+functionalized+poly(PEGMA)+brushes+on+silicon+surface+for+covalent+protein+immobilization.">NHS-ester functionalized poly(PEGMA) brushes on silicon surface for covalent protein immobilization.</a> Colloids and Surfaces B: Biointerfaces. 66 (2), 233-239 (2008).</li><li>Ma, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Facile+fabrication+of+microsphere-polymer+brush+hierarchically+three-dimensional+(3D)+substrates+for+immunoassays.">Facile fabrication of microsphere-polymer brush hierarchically three-dimensional (3D) substrates for immunoassays.</a> Chemical Communications. 51 (31), 6749-6752 (2015).</li><li>Chong, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thiocarbonylthio+compounds+[SC+(Ph)+S&#8722;+R]+in+free+radical+polymerization+with+reversible+addition-fragmentation+chain+transfer+(RAFT+Polymerization).+Role+of+the+free-radical+leaving+group+(R).">Thiocarbonylthio compounds [SC (Ph) S&#8722; R] in free radical polymerization with reversible addition-fragmentation chain transfer (RAFT Polymerization). Role of the free-radical leaving group (R).</a> Macromolecules. 36 (7), 2256-2272 (2003).</li><li>Jochum, F. D., Theato, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature-+and+Light-Responsive+Polyacrylamides+Prepared+by+a+Double+Polymer+Analogous+Reaction+of+Activated+Ester+Polymers.">Temperature- and Light-Responsive Polyacrylamides Prepared by a Double Polymer Analogous Reaction of Activated Ester Polymers.</a> Macromolecules. 42 (16), 5941-5945 (2009).</li><li>JoVE Science Education Database. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Basic+Methods+in+Cellular+and+Molecular+Biology.+The+Western+Blot.">Basic Methods in Cellular and Molecular Biology. The Western Blot.</a> JoVE. , (2018).</li><li>Chua, G. B. H., Roth, P. J., Duong, H. T. T., Davis, T. P., Lowe, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+and+Thermoresponsive+Solution+Properties+of+Poly[oligo(ethylene+glycol)+(meth)acrylamide]s:+Biocompatible+PEG+Analogues.">Synthesis and Thermoresponsive Solution Properties of Poly[oligo(ethylene glycol) (meth)acrylamide]s: Biocompatible PEG Analogues.</a> Macromolecules. 45 (3), 1362-1374 (2012).</li></ol>

The authors have nothing to disclose.

The synthesis of poly(PFPA) grafted SiO2 beads is illustrated in Figure 1. By employing APTMS as a linker molecule, poly(PFPA) brushes covalently grafted to SiO2 substrate can be prepared via a simple two-step process. Although some of the PFP units are sacrificed for the reaction with APTMS, a large number of the PFP units are expected to remain available for later reaction with either amino-PEG or antibodies. The PFP groups are known to form low energy surfaces so pol...

An erratum was issued for: <a href="https://www.jove.com/video/58843/preparation-poly-pentafluorophenyl-acrylate-functionalized-sio2-beads">Preparation of Poly(pentafluorophenyl acrylate) Functionalized SiO2 Beads for Protein Purification</a>.&#160; Throughout the article, the term &#34;3-aminopropyltriethoxysilane&#34; has been replaced with &#34;3-aminopropyltrimethoxysilane&#34;, and &#34;APTES&#34; with &#34;APTMS&#34;.

Erratum: Preparation of Poly(pentafluorophenyl acrylate) Functionalized SiO2 Beads for Protein Purification

Reactive polymer brushes have received much interest in recent years. They can be used to immobilize functional molecules on organic or inorganic materials to create activated surfaces with applications in areas such as detection and separation1,2,3,4,5. Among the reactive polymers reported, those containing pentafluorophenyl ester units are particularly useful due to their high reactivity with amines and resistance toward hydrolysis6. One such polymer is poly(PFPA), and it can be readily functionalized post-polymerization with molecules containing primary or secondary amines7,8,9,10. In one example, poly(PFPA) brushes were reacted with amino-spiropyrans to create light-responsive surfaces7.
The preparation of poly(PFPA) and its applications have been described in a number of previous publications6,7,8,9,10,11,12,13,14,15,16,17. In particular, Theato and co-workers reported the synthesis of poly(PFPA) brushes via both &#34;grafting to&#34; and &#34;grafting from&#34; methods7,8,10,11,12. In the &#34;grafting to&#34; approach, a poly(methylsilsesquioxane)-poly(pentafluorophenyl acrylate) (poly(MSSQ-PFPA)) hybrid polymer was synthesized8,10,11,12. The poly(MSSQ) component was able to form strong adhesion with a number of different organic and inorganic surfaces, thus allowing the poly(PFPA) component to form a brush layer on the coated material surface. In the &#34;grafting from&#34; approach, surface initiated reversible addition and fragmentation chain transfer (SI-RAFT) polymerization was employed to prepare poly(PFPA) brushes7. In this case, a surface immobilized chain transfer agent (SI-CTA) was first covalently attached to the substrate via silica-silane reaction. The immobilized SI-CTA then participated in the SI-RAFT polymerization of PFPA monomers, generating densely packed poly(PFPA) brushes with stable covalent linkage to the substrate.
By utilizing the poly(PFPA) brushes synthesized via SI-RAFT polymerization, we recently demonstrated the immobilization of antibodies on poly(PFPA) grafted silica particles and their subsequent application in protein purification18. The use of poly(PFPA) brushes for antibody immobilization was found to resolve a number of issues associated with current protein separation through IP. Conventional IP relies on the use of Protein A/G as a linker for antibody immobilization19,20,21. Since the use of Protein A/G allows the antibodies to be attached with a specific orientation, high target antigen recovery efficiency is achieved. However, the use of Protein A/G suffers from non-specific protein interaction as well as the loss of antibodies during protein recovery, both of which contribute to a high level of background noise. To resolve these shortcomings, direct crosslinking of the antibodies to a solid support has been explored22,23,24. The efficiency of such techniques is typically low due to the random orientation of the crosslinked antibodies. For the poly(PFPA) grafted substrate, the immobilization of antibodies is permanent, achieved through exchange reaction between PFP units and amine functionalities on antibodies. Although the antibody orientation is still random, the system benefits from having many reactive PFP sites, controllable by the degree of polymerization. Furthermore, we showed that by partial substitution of PFP units with amino-PEG, surface hydrophilicity can be tuned, further improving the protein recovery efficiency of the system18. Overall, the poly(PFPA) grafted silica particles were shown to be an effective alternative to traditional IP with reasonable efficiency as well as much cleaner background.
In this contribution, we report an alternative method to prepare poly(PFPA) grafted surface for antibody immobilization and IP application. In a simple two-step process, as illustrated in Figure 1, an APTMS linker molecule is first deposited onto the silica surface, then the poly(PFPA) polymer is covalently attached to the linker molecule through the reaction between the PFP units on the polymer and the amine functions on APTMS. This preparation method allows for the permanent crosslinking of poly(PFPA) to a substrate surface, but avoids the many complications associated with SI-CTA synthesis and SI-RAFT polymerization of poly(PFPA) brushes. Partial substitution of the PFP units with amino-PEG can still be performed, allowing fine-tuning of the polymer brush surface properties. We show the poly(PFPA) grafted silica beads thus prepared can be immobilized with antibodies and used for protein enrichment via IP. The detailed bead preparation procedure, antibody immobilization, and IP testing are documented in this article, for readers interested in seeking an alternative to conventional Protein A/G based IP.

<table>
	<tbody>
		<tr>
			<td>2,2-Azobisisobutyronitrile, 99%</td>
			<td>Daejung Chemicals</td>
			<td>1102-4405</td>
			<td></td>
		</tr>
		<tr>
			<td>Methyl alcohol for HPLC, 99.9%</td>
			<td>Duksan Pure Chemicals</td>
			<td>d62</td>
			<td></td>
		</tr>
		<tr>
			<td>Phenylmagnesium bromide solution 1.0&#160;M in THF</td>
			<td>Sigma-Aldrich</td>
			<td>331376</td>
			<td></td>
		</tr>
		<tr>
			<td>Carbon disulfide anhydrous, &#8805;99%</td>
			<td>Sigma-Aldrich</td>
			<td>335266</td>
			<td></td>
		</tr>
		<tr>
			<td>Benzyl bromide, 98%</td>
			<td>Sigma-Aldrich</td>
			<td>B17905</td>
			<td></td>
		</tr>
		<tr>
			<td>Petroleum ether, 90%</td>
			<td>Samchun Chemicals</td>
			<td>P0220</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethyl ether, 99%</td>
			<td>Daejung Chemicals</td>
			<td>4025-4404</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium sulfate anhydrous, powder,&#160;99%</td>
			<td>Daejung Chemicals</td>
			<td>5514-4405</td>
			<td></td>
		</tr>
		<tr>
			<td>Pentafluorophenyl acrylate</td>
			<td>Santa Cruz Biotechnology</td>
			<td>sc-264001</td>
			<td>contains inhibitor</td>
		</tr>
		<tr>
			<td>Aluminium oxide, activated, basic, Brockmann I</td>
			<td>Sigma-Aldrich</td>
			<td>199443</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Chloride (NaCl)</td>
			<td>Daejung Chemicals</td>
			<td>7548-4400</td>
			<td></td>
		</tr>
		<tr>
			<td>Anisole anhydrous, 99.7%</td>
			<td>Sigma-Aldrich</td>
			<td>296295</td>
			<td></td>
		</tr>
		<tr>
			<td>Silica nanoparticle</td>
			<td>Microparticles GmbH</td>
			<td>SiO2-R-0.7</td>
			<td>5% w/v aqueous suspension</td>
		</tr>
		<tr>
			<td>3-Aminopropyltrimethoxysilane, &#62;96.0%</td>
			<td>Tokyo Chemical Industry</td>
			<td>T1255</td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl sulfoxide for HPLC, &#8805;99.7%</td>
			<td>Sigma-Aldrich</td>
			<td>34869</td>
			<td></td>
		</tr>
		<tr>
			<td>Amino-terminated poly(ethylene glycol) methyl ether</td>
			<td>Polymer Source</td>
			<td>P16082-EGOCH3NH2</td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate buffered saline tablet</td>
			<td>Takara</td>
			<td>T9181</td>
			<td></td>
		</tr>
		<tr>
			<td>Tween-20</td>
			<td>Calbiochem</td>
			<td>9480</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris-HCl (pH 8.0)</td>
			<td>Invitrogen</td>
			<td>AM9855G</td>
			<td></td>
		</tr>
		<tr>
			<td>KCl</td>
			<td>Invitrogen</td>
			<td>AM9640G</td>
			<td></td>
		</tr>
		<tr>
			<td>NP-40</td>
			<td>VWR</td>
			<td>E109-50ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Glycerol</td>
			<td>Invitrogen</td>
			<td>15514-011</td>
			<td></td>
		</tr>
		<tr>
			<td>Dithiothreitol</td>
			<td>Biosesang</td>
			<td>D1037</td>
			<td></td>
		</tr>
		<tr>
			<td>Protease inhibitor</td>
			<td>Merck</td>
			<td>535140-1MLCN</td>
			<td></td>
		</tr>
		<tr>
			<td>Bromo phenol blue</td>
			<td>Sigma-Aldrich</td>
			<td>B5525-5G</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris-HCl (pH 6.8)</td>
			<td>Biosolution</td>
			<td>BT033</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium dodecyl sulfate</td>
			<td>Biosolution</td>
			<td>BS003</td>
			<td></td>
		</tr>
		<tr>
			<td>2-Mercaptoethanol</td>
			<td>Gibco</td>
			<td>21985-023</td>
			<td></td>
		</tr>
		<tr>
			<td>PKR Antibody</td>
			<td>Cell Signaling Technology</td>
			<td>12297S</td>
			<td></td>
		</tr>
		<tr>
			<td>GAPDH Antibody</td>
			<td>Santa Cruz Biotechnology</td>
			<td>sc-32233</td>
			<td></td>
		</tr>
		<tr>
			<td>Normal Rabbit IgG</td>
			<td>Cell Signaling Technology</td>
			<td>2729S</td>
			<td></td>
		</tr>
		<tr>
			<td>HeLa</td>
			<td>Korea Cell Line Bank</td>
			<td>10002</td>
			<td></td>
		</tr>
		<tr>
			<td>Sonicator</td>
			<td>DAIHAN Scientific</td>
			<td>WUC-D10H</td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrasonicator</td>
			<td>BMBio</td>
			<td>BR2006A</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge I</td>
			<td>Eppendorf</td>
			<td>5424 R</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge II</td>
			<td>LABOGENE</td>
			<td>1736R</td>
			<td></td>
		</tr>
		<tr>
			<td>Rotator</td>
			<td>FINEPCR</td>
			<td>ROTATOR/AG</td>
			<td></td>
		</tr>
		<tr>
			<td>Vacuum oven</td>
			<td>DAIHAN Scientific</td>
			<td>ThermoStable OV-30</td>
			<td></td>
		</tr>
		<tr>
			<td>Gel permeation chromatography (THF)</td>
			<td>Agilent Technologies</td>
			<td>1260 Infinity II</td>
			<td></td>
		</tr>
		<tr>
			<td>X-ray photoelectron spectrometer</td>
			<td>Thermo VG Scientific</td>
			<td>Sigma Probe</td>
			<td></td>
		</tr>
		<tr>
			<td>Dynamic light scattering</td>
			<td>Malvern Instruments</td>
			<td>ZEN 3690</td>
			<td></td>
		</tr>
	</tbody>
</table>

preparation of poly(pentafluorophenyl acrylate) functionalized sio2 beads for protein purification

We demonstrate a simple method to prepare poly(pentafluorophenyl acrylate) (poly(PFPA)) grafted silica beads for antibody immobilization and subsequent immunoprecipitation (IP) application. The poly(PFPA) grafted surface is prepared via a simple two-step process. In the first step, 3-aminopropyltrimethoxysilane (APTMS) is deposited as a linker molecule onto the silica surface. In the second step, poly(PFPA) homopolymer, synthesized via the reversible addition and fragmentation chain transfer (RAFT) polymerization, is grafted to the linker molecule through the exchange reaction between the pentafluorophenyl (PFP) units on the polymer and the amine groups on APTMS. The deposition of APTMS and poly(PFPA) on the silica particles are confirmed by X-ray photoelectron spectroscopy (XPS), as well as monitored by the particle size change measured via dynamic light scattering (DLS). To improve the surface hydrophilicity of the beads, partial substitution of poly(PFPA) with amine-functionalized poly(ethylene glycol) (amino-PEG) is also performed. The PEG-substituted poly(PFPA) grafted silica beads are then immobilized with antibodies for IP application. For demonstration, an antibody against protein kinase RNA-activated (PKR) is employed, and IP efficiency is determined by Western blotting. The analysis results show that the antibody immobilized beads can indeed be used to enrich PKR while non-specific protein interactions are minimal.

A schematic for the preparation of poly(PFPA) grafted SiO2 beads, with or without PEG substitution is shown in Figure 1. To monitor the APTMS and poly(PFPA) grafting process, bare SiO2 beads, APTMS functionalized SiO2 beads, and poly(PFPA) grafted SiO2 beads are characterized by both DLS (Figure 2) and XPS (Figure 3). IP efficiencies of the beads are determ...

Watch this Scientific Journal Video about Preparation of Polypentafluorophenyl acrylate Functionalized SiO2 Beads for Protein Purification at JoVE.com

Preparation of Polypentafluorophenyl acrylate Functionalized SiO2 Beads for Protein Purification

A protocol for the preparation of poly(pentafluorophenyl acrylate) (poly(PFPA)) grafted silica beads is presented. The poly(PFPA) functionalized surface is then immobilized with antibodies and used successfully for the protein separation through immunoprecipitation.

Preparation of Poly(pentafluorophenyl acrylate) Functionalized SiO2 Beads for Protein Purification

We demonstrate a simple method to prepare poly(pentafluorophenyl acrylate) (poly(PFPA)) grafted silica beads for antibody immobilization and subsequent ...

preparation-poly-pentafluorophenyl-acrylate-functionalized-sio2-beads

Research

JoVE Journal

Chemistry

9.5K Views.  Korea Advanced Institute of Science and Technology (KAIST). Deze methode kan worden gebruikt om biomolecuul-geactiveerde oppervlakken voor te bereiden, die toepassingen hebben op gebieden zoals de levering van geneesmiddelen, biologische doeldetectie en bioscheiding. Het belangrijkste voordeel van deze techniek is dat de poly PFPA borstels zeer reactief zijn met amines, en de immobilisatie van antilichamen wordt bereikt door incubatie in bufferoplossing voor een paar uur. De implicaties van deze techniek breiden zich uit tot de eiwitzuivering door mid...

Video: Preparation of Polypentafluorophenyl acrylate Functionalized SiO2 Beads for Protein Purification

Microwave-assisted Intramolecular Dehydrogenative Diels-Alder Reactions for the Synthesis of Functionalized Naphthalenes/Solvatochromic Dyes

Functionalized naphthalenes have applications in a variety of research fields ranging from the synthesis of natural or biologically active molecules to the preparation of new organic dyes. Although numerous strategies have been reported to access naphthalene scaffolds, many procedures still present limitations in terms of incorporating functionality, which in turn narrows the range of available substrates. The development of versatile methods for direct access to substituted naphthalenes is therefore highly desirable.
The Diels-Alder (DA) cycloaddition reaction is a powerful and attractive method for the formation of saturated and unsaturated ring systems from readily available starting materials. A new microwave-assisted intramolecular dehydrogenative DA reaction of styrenyl derivatives described herein generates a variety of functionalized cyclopenta[b]naphthalenes that could not be prepared using existing synthetic methods. When compared to conventional heating, microwave irradiation accelerates reaction rates, enhances yields, and limits the formation of undesired byproducts.
The utility of this protocol is further demonstrated by the conversion of a DA cycloadduct into a novel solvatochromic fluorescent dye via a Buchwald-Hartwig palladium-catalyzed cross-coupling reaction. Fluorescence spectroscopy, as an informative and sensitive analytical technique, plays a key role in research fields including environmental science, medicine, pharmacology, and cellular biology. Access to a variety of new organic fluorophores provided by the microwave-assisted dehydrogenative DA reaction allows for further advancement in these fields.

Microwave-assisted intramolecular dehydrogenative Diels-Alder (DA) reactions provide concise access to functionalized cyclopenta[b]naphthalene building blocks. The utility of this methodology is demonstrated by one-step conversion of the dehydrogenative DA cycloadducts into novel solvatochromic fluorescent dyes via Buchwald-Hartwig palladium-catalyzed cross-coupling reactions.

Functionalized naphthalenes have applications in a variety of research fields ranging from the synthesis of natural or biologically active molecules to ...

Synthesis and Characterization of Functionalized Metal-organic Frameworks

Metal-organic frameworks have attracted extraordinary amounts of research attention, as they are attractive candidates for numerous industrial and technological applications. Their signature property is their ultrahigh porosity, which however imparts a series of challenges when it comes to both constructing them and working with them. Securing desired MOF chemical and physical functionality by linker/node assembly into a highly porous framework of choice can pose difficulties, as less porous and more thermodynamically stable congeners (e.g., other crystalline polymorphs, catenated analogues) are often preferentially obtained by conventional synthesis methods. Once the desired product is obtained, its characterization often requires specialized techniques that address complications potentially arising from, for example, guest-molecule loss or preferential orientation of microcrystallites. Finally, accessing the large voids inside the MOFs for use in applications that involve gases can be problematic, as frameworks may be subject to collapse during removal of solvent molecules (remnants of solvothermal synthesis). In this paper, we describe synthesis and characterization methods routinely utilized in our lab either to solve or circumvent these issues. The methods include solvent-assisted linker exchange, powder X-ray diffraction in capillaries, and materials activation (cavity evacuation) by supercritical CO2 drying. Finally, we provide a protocol for determining a suitable pressure region for applying the Brunauer-Emmett-Teller analysis to nitrogen isotherms, so as to estimate surface area of MOFs with good accuracy.

Synthesis, activation, and characterization of intentionally designed metal-organic framework materials is challenging, especially when building blocks are incompatible or unwanted polymorphs are thermodynamically favored over desired forms. We describe how applications of solvent-assisted linker exchange, powder X-ray diffraction in capillaries and activation via supercritical CO2 drying, can address some of these challenges.

Metal-organic frameworks have attracted extraordinary amounts of research attention, as they are attractive candidates for numerous industrial and ...

Supercritical Nitrogen Processing for the Purification of Reactive Porous Materials

Supercritical fluid extraction and drying methods are well established in numerous applications for the synthesis and processing of porous materials. Herein, nitrogen is presented as a novel supercritical drying fluid for specialized applications such as in the processing of reactive porous materials, where carbon dioxide and other fluids are not appropriate due to their higher chemical reactivity. Nitrogen exhibits similar physical properties in the near-critical region of its phase diagram as compared to carbon dioxide: a widely tunable density up to ~1 g ml-1, modest critical pressure (3.4 MPa), and small molecular diameter of ~3.6 &#197;. The key to achieving a high solvation power of nitrogen is to apply a processing temperature in the range of 80-150 K, where the density of nitrogen is an order of magnitude higher than at similar pressures near ambient temperature. The detailed solvation properties of nitrogen, and especially its selectivity, across a wide range of common target species of extraction still require further investigation. Herein we describe a protocol for the supercritical nitrogen processing of porous magnesium borohydride.

Nitrogen is an effective supercritical fluid for extraction or drying processes due to its small molecular size, high density in the near-liquid supercritical regime, and chemical inertness. We present a supercritical nitrogen drying protocol for the purification treatment of reactive, porous materials.

Supercritical fluid extraction and drying methods are well established in numerous applications for the synthesis and processing of porous materials. ...

Microbubble Fabrication of Concave-porosity PDMS Beads

Microbubble fabrication (by use of a fine emulsion) provides a means of increasing the surface-area-to-volume (SAV) ratio of polymer materials, which is particularly useful for separations applications. Porous polydimethylsiloxane (PDMS) beads can be produced by heat-curing such an emulsion, allowing the interface between the aqueous and aliphatic phases to mold the morphology of the polymer. In the procedures described here, both polymer and crosslinker (triethoxysilane) are sonicated together in a cold-bath sonicator. Following a period of cross-linking, emulsions are added dropwise to a hot surfactant solution, allowing the aqueous phase of the emulsion to separate, and forming porous polymer beads. We demonstrate that this method can be tuned, and the SAV ratio optimized, by adjusting the electrolyte content of the aqueous phase in the emulsion. Beads produced in this way are imaged with scanning electron microscopy, and representative SAV ratios are determined using Brunauer&#8211;Emmett&#8211;Teller (BET) analysis. Considerable variability with the electrolyte identity is observed, but the general trend is consistent: there is a maximum in SAV obtained at a specific concentration, after which porosity decreases markedly.

Procedures used to generate microstructured concave-porosity polydimethylsiloxane beads are presented. Effects of electrolyte concentration and identity within the aqueous phase are particularly emphasized.

Microbubble fabrication (by use of a fine emulsion) provides a means of increasing the surface-area-to-volume (SAV) ratio of polymer materials, which is ...

Synthesis of Programmable Main-chain Liquid-crystalline Elastomers Using a Two-stage Thiol-acrylate Reaction

This study presents a novel two-stage thiol-acrylate Michael addition-photopolymerization (TAMAP) reaction to prepare main-chain liquid-crystalline elastomers (LCEs) with facile control over network structure and programming of an aligned monodomain. Tailored LCE networks were synthesized using routine mixing of commercially available starting materials and pouring monomer solutions into molds to cure. An initial polydomain LCE network is formed via a self-limiting thiol-acrylate Michael-addition reaction. Strain-to-failure and glass transition behavior were investigated as a function of crosslinking monomer, pentaerythritol tetrakis(3-mercaptopropionate) (PETMP). An example non-stoichiometric system of 15 mol% PETMP thiol groups and an excess of 15 mol% acrylate groups was used to demonstrate the robust nature of the material. The LCE formed an aligned and transparent monodomain when stretched, with a maximum failure strain over 600%. Stretched LCE samples were able to demonstrate both stress-driven thermal actuation when held under a constant bias stress or the shape-memory effect when stretched and unloaded. A permanently programmed monodomain was achieved via a second-stage photopolymerization reaction of the excess acrylate groups when the sample was in the stretched state. LCE samples were photo-cured and programmed at 100%, 200%, 300%, and 400% strain, with all samples demonstrating over 90% shape fixity when unloaded. The magnitude of total stress-free actuation increased from 35% to 115% with increased programming strain. Overall, the two-stage TAMAP methodology is presented as a powerful tool to prepare main-chain LCE systems and explore structure-property-performance relationships in these fascinating stimuli-sensitive materials.

A novel methodology is presented to synthesize and program main-chain liquid-crystalline elastomers using commercially available starting monomers. A wide range of thermomechanical properties was tailored by adjusting the amount of crosslinker, while the actuation performance was dependent on the amount of applied strain during programming.

This study presents a novel two-stage thiol-acrylate Michael addition-photopolymerization (TAMAP) reaction to prepare main-chain liquid-crystalline ...

Immobilization of Multi-biocatalysts in Alginate Beads for Cofactor Regeneration and Improved Reusability

We have recently developed a simple, reusable and coupled whole-cell biocatalytic system with the capability of cofactor regeneration and biocatalyst immobilization for improved production yield and sustained synthesis. Described herewith is the experimental procedure for the development of such a system consisting of two E. coli strains that express functionally complementary enzymes. Together, these two enzymes can function co-operatively to mediate the regeneration of expensive cofactors for improving the product yield of the bioreaction. In addition, the method of synthesizing an immobilized form of the coupled biocatalytic system by encapsulation of whole cells in calcium alginate beads is reported. As an example, we present the improved biosynthesis of L-xylulose from L-arabinitol by coupling E. coli cells expressing the enzymes L-arabinitol dehydrogenase or NADH oxidase. Under optimal conditions and using an initial concentration of 150 mM L-arabinitol, the maximal L-xylulose yield reached 96%, which is higher than those reported in the literature. The immobilized form of the coupled whole-cell biocatalysts demonstrated good operational stability, maintaining 65% of the yield obtained in the first cycle after 7 cycles of successive re-use, while the free cell system almost completely lost the catalytic activity. Therefore, the methods reported here provides two strategies that could help improve the industrial production of L-xylulose, as well as other value-added compounds requiring the use of cofactors in general.

Presented is the protocol for co-immobilizing whole-cell biocatalysts for cofactor regeneration and improved reusability, using the production of L-xylulose as an example. The cofactor regeneration is achieved by coupling two Escherichia coli strains expressing functionally complementary enzymes; the whole-cell biocatalyst immobilization is achieved by cell encapsulation in calcium alginate beads.

We have recently developed a simple, reusable and coupled whole-cell biocatalytic system with the capability of cofactor regeneration and biocatalyst ...

Preparation of a Corannulene-functionalized Hexahelicene by Copper(I)-catalyzed Alkyne-azide Cycloaddition of Nonplanar Polyaromatic Units

The main purpose of this video is to show 6 reaction steps of a convergent synthesis and prepare a complex molecule containing up to three nonplanar polyaromatic units, which are two corannulene moieties and a racemic hexahelicene linking them. The compound described in this work is a good host for fullerenes. Several common organic reactions, such as free-radical reactions, C-C coupling or click chemistry, are employed demonstrating the versatility of functionalization that this compound can accept. All of these reactions work for planar aromatic molecules. With subtle modifications, it is possible to achieve similar results for nonplanar polyaromatic compounds.

Preparation of a Corannulene-functionalized Hexahelicene by CopperI-catalyzed Alkyne-azide Cycloaddition of Nonplanar Polyaromatic Units

Here, we present a protocol to synthesize a complex organic compound comprised of three nonplanar polyaromatic units, assembled easily with reasonable yields.

The main purpose of this video is to show 6 reaction steps of a convergent synthesis and prepare a complex molecule containing up to three nonplanar ...

The Effect of Interfacial Chemical Bonding in TiO2-SiO2 Composites on Their Photocatalytic NOx Abatement Performance

The chemical bonding of particulate photocatalysts to supporting material surfaces is of great importance in engineering more efficient and practical photocatalytic structures. However, the influence of such chemical bonding on the optical and surface properties of the photocatalyst and thus its photocatalytic activity/reaction selectivity behavior has not been systematically studied. In this investigation, TiO2 has been supported on the surface of SiO2 by means of two different methods: (i) by the in situ formation of TiO2 in the presence of sand quartz via a sol-gel method employing tetrabutyl orthotitanium (TBOT); and (ii) by binding the commercial TiO2 powder to quartz on a surface silica gel layer formed from the reaction of quartz with tetraethylorthosilicate (TEOS). For comparison, TiO2 nanoparticles were also deposited on the surfaces of a more reactive SiO2 prepared by a hydrolysis-controlled sol-gel technique as well as through a sol-gel route from TiO2 and SiO2 precursors. The combination of TiO2 and SiO2, through interfacial Ti-O-Si bonds, was confirmed by FTIR spectroscopy and the photocatalytic activities of the obtained composites were tested for photocatalytic degradation of NO according to the ISO standard method (ISO 22197&#8722;1). The electron microscope images of the obtained materials showed that variable photocatalyst coverage of the support surface can successfully be achieved but the photocatalytic activity towards NO removal was found to be affected by the preparation method and the nitrate selectivity is adversely affected by Ti-O-Si bonding.

The Effect of Interfacial Chemical Bonding in TiO2-SiO2 Composites on Their Photocatalytic NOx Abatement Performance

The focus of the present work is to establish means to generate and quantify levels of Ti-O-Si linkages and to correlate these with the photocatalytic properties of the supported TiO2.

The chemical bonding of particulate photocatalysts to supporting material surfaces is of great importance in engineering more efficient and practical ...

Methionine Functionalized Biocompatible Block Copolymers for Targeted Plasmid DNA Delivery

Reversible addition-fragmentation chain transfer (RAFT) polymerization integrates the advantages of radical polymerization and living polymerization. This work presents the preparation of methionine functionalized biocompatible block copolymers via RAFT polymerization. Firstly, N,N-bis(2-hydroxyethyl)methacrylamide-b-N-(3-aminopropyl)methacrylamide (BNHEMA-b-APMA, BA) was synthesized via RAFT polymerization using 4,4&#8217;-azobis(4-cyanovaleric acid) (ACVA) as an initiating agent and 4-cyanopentanoic acid dithiobenzoate (CTP) as the chain transfer agent. Subsequently, N,N-bis(2-hydroxyethyl)methacrylamide-b-N-(3-guanidinopropyl)methacrylamide (methionine grafted BNHEMA-b-GPMA, mBG) was prepared by modifying amine groups in APMA with methionine and guanidine groups. Three kinds of block polymers, mBG1, mBG2, and mBG3, were synthesized for comparison. A ninhydrin reaction was used to quantify the APMA content; mBG1, mBG2, and mBG3 had 21%, 37%, and 52% of APMA, respectively. Gel permeation chromatography (GPC) results showed that BA copolymers possess molecular weights of 16,200 (BA1), 20,900(BA2), and 27,200(BA3) g/mol. The plasmid DNA (pDNA) complexing ability of the obtained block copolymer gene carriers was also investigated. The charge ratios (N/P) were 8, 16, and 4 when pDNA was complexed completely with mBG1, mBG2, mBG3, respectively. When the N/P ratio of mBG/pDNA polyplexes was higher than 1, the Zeta potential of mBG was positive. At an N/P ratio between 16 and 32, the average particle size of mBG/pDNA polyplexes was between 100-200 nm. Overall, this work illustrates a simple and convenient protocol for the block copolymer carrier synthesis.

This work presents the preparation of methionine functionalized biocompatible block copolymers (mBG) via the reversible addition-fragmentation chain transfer (RAFT) method. The plasmid DNA complexing ability of the obtained mBG and their transfection efficiency were also investigated. The RAFT method is very beneficial for polymerizing monomers containing special functional groups.

Reversible addition-fragmentation chain transfer (RAFT) polymerization integrates the advantages of radical polymerization and living polymerization. This ...

Efficient Synthesis of All-Carbon Quaternary Centers via the Conjugate Addition of Functionalized Monoorganozinc Bromides

The conjugate addition of organometallic reagents to &#945;,&#946;-unsaturated carbonyls represents an important method to generate C&#8211;C bonds in the preparation of all-carbon quaternary centers. Though conjugate additions of organometallic reagents are typically performed utilizing highly reactive organolithium or Grignard reagents, organozinc reagents have garnered attention for their enhanced chemoselectivity and mild reactivity. Despite numerous recent advances with more reactive diorganozinc and mixed diorganozinc reagents, the generation of all-carbon quaternary centers via the conjugate addition of functionalized monoorganozinc reagents remains a challenge. This protocol details a convenient and mild &#8220;one-pot&#8221; preparation and copper mediated conjugate addition of functionalized monoorganozinc bromides to cyclic &#945;,&#946;-unsaturated carbonyls to afford a broad scope of all-carbon quaternary centers in generally excellent yield and diastereoselectivity. Key to the development of this technology is the utilization of DMA as a reaction solvent with TMSCl as a Lewis acid. Notable advantages to this methodology include the operational simplicity of the organozinc reagent preparation afforded by the utilization of DMA as a solvent, as well as an efficient conjugate addition mediated by various Cu(I) and Cu(II) salts. Moreover, an intermediate silyl enol ether can be isolated utilizing a modified workup procedure. The substrate scope is limited to cyclic unsaturated ketones, and the conjugate addition is impeded by stabilized (e.g., allyl, enolate, homoenolate) and sterically encumbered (e.g., neopentyl, o-aryl) monoorganozinc reagents. Conjugate additions to five- and seven-membered rings were effective, albeit in lower yields compared with six-membered ring substrates.

A simple and practical protocol for the efficient conjugate addition of functionalized monoorganozinc bromides to cyclic &#945;,&#946;-unsaturated carbonyls to furnish all-carbon quaternary centers was developed.

The conjugate addition of organometallic reagents to &#945;,&#946;-unsaturated carbonyls represents an important method to generate C&#8211;C bonds in the ...

Preparation of Poly(pentafluorophenyl acrylate) Functionalized SiO₂ Beads for Protein Purification

Samenvatting

Meer video's verkennen

Hoofdstukken in deze video