sodium;dodecyl sulfate

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Western blotting was performed with the compound of the example 64 of the present invention. A375P cell line was lysed by using RIPA buffer solution (50 mM tris pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40, 0.25% sodium deoxycholate) containing phosphatase inhibitor (Phosphatase inhibitor cocktail I & II) and protease inhibitor (Complete, Mini, EDTA-free; Roche Applied Science), and centrifuged at 13000 rpm for 10 min. Whole protein was stained with Bio-Rad agent (bio-rad #500-0006) and quantitated by comparing with BSA (positive control). 12% gel was prepared by using 30% acrylamide, 1.5 M Tris (pH 8.8), 10% SDS solution, 10% PAS solution and TEMED. After loading 100 ug of protein per one lane of gel, proteins were isolated by sodiumdodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE, 250 mM Tris base, 2.5 M Glycine, 0.1% SDS, 140V, electrophoresis for 2 h), and transferred to nitrocellulose filter at 200 mA for 2 h. The results are weakly reacted with an antibody[Rabbit anti-Map Kinase (Zymed #61-7400) and phosphor-p44/42 MAP kinase (Thr202/Tyr204) (Cell signaling #9101)] at 4° C. for a day. Further, it is washed with TBST buffer solution containing 0.05% tween20 at 4 times every 5 minutes. The results are weakly reacted with secondary antibody[anti-rabbit IgG (Amersham #NA934V)] for one hour at room temperature, and washed by the same manner. Protein bands were identified with imaging apparatus (Gelliance). The results is shown in FIG. 1. 12% gel was prepared

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Bacterial cells were harvested by centrifugation and resuspended in SDS sample buffer (0.0625M Tris, pH 6.8, 10% glycerol, 5% mercaptoethanol, 2.3% SDS). The resuspended pellet was boiled for 5 min. and then cleared of insoluble cellular debris by centrifugation. The supernatants obtained from IPTG-induced cultures of GE3-2, GE9-2, GE15-1, GE17-2, GE4-8, EXP3-7, GE1-N and GE57 were analyzed by SDS-polyacrylamide gel electrophoresis (PAGE) together with uninduced lysates. The proteins from these gels were then transferred to nitrocellulose filters (i.e., by Western blotting).

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: When sodium lauryl sulfate is used as the surfactant (4), the acid of step (B) is preferably a dilute mineral acid, such as hydrochloric acid. The dilute hydrochloric acid reacts with the sodium lauryl sulfate to give hydrogen lauryl sulfate, which is an active polymerization catalyst for the polydiorganosiloxane oligomer.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Same as C5 except that the monomer emulsion was composed of 5 g of SDS, 1 g of sodium persulfate, 80 g of ethyl methacrylate, 100 g of n-butyl methacrylate, 10 g of methacrylic acid, and 10 g of ethylene glycol dimethacrylate. Glass transition temperature was 69° C., average final particle size was 39 nm and % solid was 20.9%.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Same as C5 except that the monomer emulsion was composed of 5 g of SDS, 1 g of sodium persulfate, 90 g of ethyl methacrylate, 90 g of n-butyl methacrylate, 10 g of methacrylic acid, and 10 g of ethylene glycol dimethacrylate. Glass transition temperature was 66° C., average particle size was 38 nm and % solids was 21.1%.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Same as C5 except that the monomer emulsion was composed of 5 g of SDS, 1 g of sodium persulfate, 60 g of ethyl methacrylate, 100 g of n-butyl methacrylate, 20 g of methacrylic acid, and 20 g of ethylene glycol dimethacrylate. The final particle size was 34 nm, % solids was 21.1% and Tg was 89° C.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Same as C5 except that monomer emulsion was composed of 5 g of SDS, 1 g of sodium persulfate, 110 g of ethyl methacrylate, 70 g of n-butyl methacrylate, and 20 g of ethylene glycol dimethacrylate. Glass transition temperature was 60° C., average particle size was 29 nm and % solids was 20.7%.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Same as C5 except that monomer emulsion was composed of 5 g of SDS, 1 g of sodium persulfate, 180 g of ethyl methacrylate, and 20 g of ethylene glycol dimethacrylate. Tg was 74° C., average particle size was 33 nm and % solids was 20.4%.

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Nuclei were isolated from suspension cultured cells according to the procedure of Lawton and Lamb, Mol. Cell Biol. 7, 335-341 (1987). Isolated nuclei were incubated in 0.1M ammonium sulfate, 4 mM MgCl2, 0.3 mM phosphocreatine, 0.15 mg/ml creatine phosphokinase, 1 U/ml RNasin (Promega), 0.5 mL each ATP, CTP, GTP, and 2 mCi/ml 32P-UTP (3000 Ci/mmol). The standard reaction volume was 30 ml containing 15 ml nuclei. The reaction mixture was incubated 30 minutes at 30° C. Three units RQ1 DNAse (Promega) and 2 ml 20 mM CaCl2 were added and the reaction mix incubated at 26° C. for 5 minutes. A mixture containing 1.5 ml proteinase K (2 mg/ml), 9 ml 10×SET (1×SET is 0.5% SDS, 5 mM EDTA, 10 mM Tris-HCl pH 7.4) and 50 mg yeast tRNA was added and the reaction was continued at 37° C. for 30 minutes. Labelled RNA was isolated by acid guanidinium/phenol/chloroform extraction. The assay consistently yielded 2-5×106CPMs. Nuclei were isolated from suspension cultured cells containing 15 ml the reaction mix incubated at 26° C. for 5 minutes was added the reaction was continued at 37° C. for 30 minutes

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Bacterial cells were harvested by centrifugation and resuspended in SDS sample buffer (0.0625M Tris, pH 6.8, 10% glycerol, 5% mercaptoethanol, 2.3% SDS). The resuspended pellet was boiled for 5 min. and then cleared of insoluble cellular debris by centrifugation. The supernatants obtained from IPTG-induced cultures of GE3-2, GE9-2, GE15-1, GE17-2, GE4-8, EXP3-7, GE1-N and GE57 were analyzed by SDS-polyacrylamide gel electrophoresis (PAGE) together with uninduced lysates. The proteins from these gels were then transferred to nitrocellulose filters (i.e., by Western blotting).

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: More specifically, cultured J774 cells were plated on a 24F culture plate containing a serum-free RPMI 1640 medium so as to account for 1×106 cells/well. Added to the cells were [14C]oleic acid, delipidized BAS (bovine serum albumin), reconstituted ribosome (0.3M glucose solution containing cholesterol and phosphatidylcholine at a weight ratio of 2:1) and 25-hydroxycholesterol to culture the cells for 4 hours at 37° C. under 5% CO2. After the culturing, the cultured cells were disrupted with a 1% SDS (sodium lauryl sulfate) solution, and the lipid in the disrupted solution was extracted with hexane. The extract was dried to solid under reduced pressure. After the resultant residue was then developed (developing solvent: diethyl ether/hexane/acetic acid=80/20/1) by TLC (thin-layer chromatography), the amount of cholesteryl oleate formed was determined by an imaging plate. containing a serum-free RPMI 1640 medium so as Added to the cells the lipid in the disrupted solution was extracted with hexane The extract was dried to solid under reduced pressure

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution was prepared of 6.0 g of sodium dodecylsulfate in 300 mL of deoxygenated water. N-Hexadecylacrylamide, 0.666 g, was dissolved in this solution followed by 9.44 g of acrylamide, 8.92 g of DMA, 10.00 g of N-vinylpyrrolidinone (NVP) and 27.99 g of sodium acrylamido-2-methylpropane sulfonate. The resulting solution was carefully transferred to a 1 L Morton style resin kettle fitted with a chilled water condenser, thermometer, inert gas sparger and mechanical stirrer. The temperature was adjusted to 25 C. and polymerization was initiated by the addition of 0.0518 g of 2,2'-azobis(N,N'-dimethylene-isobutyramidine) dihydrochloride. After stirring for 16 hr at 25 ±0.2 C., a 250 mL portion of the viscous solution was added to 3 L of acetone. The precipitated polymer was then masticated in a Waring blendor with methanol/acetone, filtered and dried under vacuum at 30 C. The yield of polymer was 42.21 g. fitted with a chilled water condenser and polymerization filtered dried under vacuum at 30 C