Difference between revisions of "20.109(F07): Phage by design, pt2"
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{{Template:20.109(F07)}} | {{Template:20.109(F07)}} | ||
==Introduction== | ==Introduction== | ||
− | [[Image: | + | Electrodeposition is a commonly used technique in which current is applied to layer one material, usually a metal, onto another.[[Image:200px-1937-Wheat-Penny-Front-Back.jpg|thumb|left|Wikipedia's image of an all-copper penny from 1937]] Modern pennies may be the most pervasive example of electrodeposition though older ones tucked away in piggy banks are not. Pennies used to be made of pure or nearly pure copper, but in 1982 when the price of copper increased and an all-copper penny had more than a penny s worth of copper in it, a new kind of cent coin was minted. Post-1982 pennies are made with an inexpensive metal (zinc) that is covered with a thin copper-coat. [[Image:EarFlaresWithCondors giltcopper.jpg|thumb|right|Pair of earflares with condors from Loma Negra, Pirua Valley, 2nd-3rd century, The Metropolitan Museum of Art photo credit:web.site@metmuseum.org.]]The coating is applied using the positive terminal of a power supply to oxidize the material on the anode (a lump of copper in the case of pennies), allowing it to react with anions in solution. At the cathode, the material gets reduced (i.e. gains electrons), regains its zero valence state and is deposited on the cathode surface (i.e. zinc if we re still talking pennies). The electrolytes solution that carries the anode material to the cathode is often made from very nasty chemicals like cyanides since these are great conductors and prevent corrosion of the anode. An active search for friendlier electrolytes continues, as does optimization of electroless deposition. Even without batteries, which Volta invented in the 1800s, incredible artifacts with decorative patterns of metal are found among some ancient ruins ([http://www.lablaa.org/blaavirtual/publicacionesbanrep/bolmuseo/1996/jldi41/jldi08a.htm e.g. in Loma Negra Peru]). Replacement plating was used as much as [http://www.jstor.org/view/00778958/ap050032/05a00080/0 2000 years ago to inlay the reactive element copper with less reactive silver and gold]. |
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+ | Successful electrodeposition requires that the cathode surface be extremely clean. Hydrophobic oils only molecules thick can inhibit deposition and cathode surfaces must be extensively cleaned with solvents, detergents, sonication (which uses sound waves to remove dirt) and/or acidic baths. Technical standards for cleaning, testing and assessing electrodeposited materials are described by [http://www.astm.org/cgi-bin/SoftCart.exe/index.shtml?L+mystore+hoqt3793+1196134766 the American Society for Testing and Materials]. Their guide for cleaning metals prior to electroplating has directions for precleaning, intermediate and final-cleaning as well as trouble shooting! [http://www.astm.org/cgi-bin/SoftCart.exe/DATABASE.CART/REDLINE_PAGES/B322.htm?L+mystore+hoqt3793+1196136492]. | ||
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+ | [[Image:ITOphage tangle.png]] Today you will start to assemble your electrochromic device by electrodepositing IrO2 nanowires that formed on the specialize M13 phage. The IrO2 wires have a slight negative charge. Consequently they can be directed toward the patterned ITO slide if the slide is attached to the positive terminal of a power supply. The circuit is completed by attaching a platinum mesh to the ground terminal of a power supply. After this deposition, you will have a tangle of nanowires adhering to the areas of your slide that were not etched away last time. Next time you will finish building the electrochromic device, adding the Li+-layer of electrolytes and a second, unpatterned ITO slide to serve as the other conductive surface. A preview of the assembled ECD is found [http://openwetware.org/wiki/Image:ECD_off.png here] | ||
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==Protocols== | ==Protocols== | ||
===Part 1: M13.1 titers=== | ===Part 1: M13.1 titers=== | ||
− | The strains you transformed last time with M13K07 and M13.1 were inoculated into LB+Kan and grown overnight. If the two forms of the phage are equally robust, then the phage titer in the liquid surrounding the cells should be approximately the same. Ditto if the wild type strain, MG1655, and the MDS strain are equally capable of phage production. You will compare the PFU/ul of each culture to determine if this is true. The titering protocol is identical to the one from [ | + | The strains you transformed last time with M13K07 and M13.1 were inoculated into LB+Kan and grown overnight. If the two forms of the phage are equally robust, then the phage titer in the liquid surrounding the cells should be approximately the same. Ditto if the wild type strain, MG1655, and the MDS strain are equally capable of phage production. You will compare the PFU/ul of each culture to determine if this is true. The titering protocol is identical to the one from [http://openwetware.org/wiki/20.109(F07):_Agarose_gel_electrophoresis #Protocols| module 1]. Since you don't know if the phage production will be improved or diminished by any of the variations, you will titer a wider-than-usual range of concentrations. |
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===Part 2: Phage deposition onto patterned slide=== | ===Part 2: Phage deposition onto patterned slide=== | ||
− | + | [[Image:Deposition set up.JPG|thumb|right|200px|How to set up the ringstand for the deposition of the nanowires onto the ITO slide.]] | |
#Clean the ITO slide by placing it into a 50 ml falcon tube with 1% Liquinox. Sonicate for 2 minutes. Remove the slide from the Liquinox solution with gloves or with tweezers and briefly rinse in Millipore H2O. Place the slide in a 50 mL Falcon tube with methanol. Sonicate for 2 minutes. Remove the slide and then let it air dry on a paper towel on the bench. Do not wipe the ITO surface. | #Clean the ITO slide by placing it into a 50 ml falcon tube with 1% Liquinox. Sonicate for 2 minutes. Remove the slide from the Liquinox solution with gloves or with tweezers and briefly rinse in Millipore H2O. Place the slide in a 50 mL Falcon tube with methanol. Sonicate for 2 minutes. Remove the slide and then let it air dry on a paper towel on the bench. Do not wipe the ITO surface. | ||
#Plasma clean the slide for 5 minutes. One of the teaching faculty will show you how to use the plasma cleaner. | #Plasma clean the slide for 5 minutes. One of the teaching faculty will show you how to use the plasma cleaner. | ||
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#Fill the beaker with 45 mL of Millipore H2O. Add 5 mL of the nanowire solution and mix with a pipette. | #Fill the beaker with 45 mL of Millipore H2O. Add 5 mL of the nanowire solution and mix with a pipette. | ||
#Lower the platinum mesh into the solution. Try to get as much of the platinum into the solution WITHOUT letting the tweezers touch the solution. | #Lower the platinum mesh into the solution. Try to get as much of the platinum into the solution WITHOUT letting the tweezers touch the solution. | ||
− | #Lower the ITO slide into the solution with the ITO side facing the platinum. Make sure that your pattern is fully immersed in the solution (or else you | + | #Lower the ITO slide into the solution with the ITO side facing the platinum. Make sure that your pattern is fully immersed in the solution (or else you won t get deposition on your entire pattern), but do not let the tweezers touch the solution. [[Image:ECD_set_up_close_up.JPG|thumb|left|200px|Your platinum mesh and ITO slide should be parallel to each other.]] |
− | #The platinum electrode should be about | + | #The platinum electrode should be about |