Acrylamide reagents

Citrate buffer and saline (MCB) solutions were preelectrolyzed for 24 h at — 1100 mV versus SCE to remove metal ion impurities. Acetonitrile (MCB) was redistilled and stored over molecular sieves. DTPA (Aldrich), triethylamine (Aldrich), and isobutylchloroformate (Sigma) were of the highest available purity. HSA (essentially globulin free) and rabbit IgG specific for HSA (Sigma) were obtained in the lyophilized form. Indium was obtained as anhydrous, ultrapure InCls (Alfa Products). All solutions were prepared from distilled/deionized HgO of at least 10 ft resistivity. Protein A from Staphylococcus aureus (SPA, Enzyme Center) was employed as an immunoadsorbent. It was obtained as a 10% (w/v) suspension with a 1.47 mg/ml binding capacity for immune complexes. 2-mercaptoethanol, sodium dodecyl sulfate, and acrylamide reagents were all of electrophoretic grade (Bio-Rad). 

Acrylamide, C H NO, is an interesting difiinctional monomer containing a reactive electron-deficient double bond and an amide group, and it undergoes reactions typical of those two functionalities. It exhibits both weak acidic and basic properties. The electron withdrawing carboxamide group activates the double bond, which consequendy reacts readily with nucleophilic reagents, eg, by addition. 

Starch is a polysaccharide found in many plant species. Com and potatoes are two common sources of industrial starch. The composition of starch varies somewhat in terms of the amount of branching of the polymer chains (11). Its principal use as a flocculant is in the Bayer process for extracting aluminum from bauxite ore. The digestion of bauxite in sodium hydroxide solution produces a suspension of finely divided iron minerals and siUcates, called red mud, in a highly alkaline Hquor. Starch is used to settle the red mud so that relatively pure alumina can be produced from the clarified Hquor. It has been largely replaced by acryHc acid and acrylamide-based (11,12) polymers, although a number of plants stiH add some starch in addition to synthetic polymers to reduce the level of residual suspended soHds in the Hquor. Starch [9005-25-8] can be modified with various reagents to produce semisynthetic polymers. The principal one of these is cationic starch, which is used as a retention aid in paper production as a component of a dual system (13,14) or a microparticle system (15). 

The most commonly used combination of chemicals to produce a polyacrylamide gel is acrylamide, bis acrylamide, buffer, ammonium persulfate, and tetramethylenediarnine (TEMED). TEMED and ammonium persulfate are catalysts to the polymerization reaction. The TEMED causes the persulfate to produce free radicals, causing polymerization. Because this is a free-radical driven reaction, the mixture of reagents must be degassed before it is used. The mixture polymerizes quickly after TEMED addition, so it should be poured into the gel-casting apparatus as quickly as possible. Once the gel is poured into a prepared form, a comb can be appHed to the top portion of the gel before polymerization occurs. This comb sets small indentations permanently into the top portion of the gel which can be used to load samples. If the comb is used, samples are then typically mixed with a heavier solution, such as glycerol, before the sample is appHed to the gel, to prevent the sample from dispersing into the reservoir buffer. 

Both the ease of use of this method for characterization of proteins and nucleic acids, and the abiHty to analyze many samples simultaneously for comparative purposes, have led to the prevalence of this technique. The drawbacks of a polyacrylamide matrix is that acrylamide is a neurotoxin, the reagents must be combined extremely carefiiUy, and the gels are not as pHable as most agarose gels. 

Typically, an acetanilide (1 mol. equiv.) was treated with the Vilsmeier reagent generated from POCI3 (7 mol. equiv.) and V,V-dimethylformamide (DMF, 2.5 mol. equiv.) at 75 °C for 4 - 20 h. The reaction products were readily obtained by filtration after pouring the reaction mixture onto ice-water minor reaction products were isolated after basification of the filtrate. A variety of acetanilides were studied under these optimised reaction conditions and some significant observations were noted. Activated acetanilides 3 [e.g. R = 4-Me (70%), 4-OMe (56%)] reacted faster and in better yield to give quinolines 4 than other strongly deactivated systems 3 [e.g. R = 4-Br (23%), 4-Cl (2%), 4-NO2 (0%)] — in these cases, formamidines 5 and acrylamides 6 were the major reaction products. 

Other reasons for a wide propagation of polymerization in water include (1) reduction of energy consumed to separate the initial monomer in crystal form (acrylamide is produced and used in the aqueous solution form), which, in addition, is associated with the probability of its spontaneous polymerization, and (2) recovery of the organic solvents, which results in less environmental pollution and the elimination of the stage of solution of polymer reagents used, as a rule, in the form of the aqueous solutions. 

Haas) and I(-( hydroxymethyl)acrylamlde (IJ-methylol acrylamide, American Cyanamld) were used without purification. Acetonitrile was dried over molecular sieves (4A) and distilled prior to use. Ethanolamlne and l-amlno-2-propanol were vacuum distilled. Reagent grade acryloyl chloride, 2-propanol, and sodium persulfate were used as received. 

Polj/meric supports based on polyacrylamide are s)mthesized by copol3tmerization of acrylamide and a cross-linking reagent and can be used directly in affinity chromatography due to its more hydrophilic properties than polystyrene supports. Polyacrylamide gels are either soft... 

TABLE 1. Copolymers of Trans-l,4-bromo-2-butene Formed by Condensing with Selected nucleophiles in AyV-dimethyl Acrylamide Using a 1 1 Reagent Charge"... 

Perhaps the most difficult and inconvenient aspect of polyacrylamide gel electrophoresis is the preparation of gels. The monomer, acrylamide, is a neurotoxin and a cancer suspect agent hence, special handling is required. Other necessary reagents including catalysts and initiators also require special handling and are unstable. In addition, it is difficult to make gels that have reproducible thicknesses and compositions. Many researchers are now... 

To prepare a 12% acrylamide gel, mix the following reagents and solutions in a 50-mL Erlenmeyer flask ... 

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