Chemi-Lumi One L
Contents
1. TBS T or PBS T Dilution buffer Use one of the followings a TBS T or PBS T b 5 Blocking One in TBS T or PBS T recommended for reducing high background C Signal Enhancer HIKARI Product No 02770 recommended for increasing weak signal 1 2 For research use only Manufactured by NACALAI TESQUE INC not intended for diagnostic or clinical use 498 Higashitamaya cho Nijo Karasuma Nakagyo ku Kyoto 604 0855 Japan R Contact us nacalai tesque Cf Email info intl nacalai co jp The quality for certainty Y TEL gt 81 75 251 1730 CF FAX 81 75 251 1763 07880E_1303_5 Protocol Following protocol is an example Modify accordingly if necessary 1 Transfer proteins to membrane from electrophoresis gel and wash the membrane with Washing buffer for 5 minutes Repeat twice Block the membrane with Blocking buffer Wash the membrane with Washing buffer for 5 minutes Repeat twice Incubate the membrane with a primary antibody at room temperature for 1 hour Wash the membrane with Washing buffer for 5 minutes Repeat twice Incubate the membrane with a HRP conjugated secondary antibody at room temperature for 1 hour Wash the membrane with Washing buffer for 5 minutes Repeat twice Prepare the Working solution Mix Solution A and Solution B in one to one ratio Required volume is 0 125ml cm membrane Remove Washing buffer from the membrane carefully with paper towels 10 Spread a plastic wrap on th2. 80 70 Solution A 250ml x 1 Solution B 250ml x 1 for 4 000 cm 2x50mI Product No 07880 54 Solution A 50ml x1 Solution B 50ml x 1 for 800 cm 2 2 For research use only Manufactured by NACALAI TESQUE INC not intended for diagnostic or clinical use 498 Higashitamaya cho Nijo Karasuma Nakagyo ku Kyoto 604 0855 Japan
3. R Contact us nacalai tesque Cf Email info intl nacalai co jp l Y TEL 81 75 251 1730 FAX 81 75 251 1763 07880E_1303_5 The quality for certainty Product No 07880 Chemi Lumi One L Features Chemi Lumi One is a series of high sensitive luminol based chemiluminescence assay kits for western blotting Chemi lumi One L provides picogram detection of proteins with long signal duration in western blot applications Chemi Lumi One L Chemi Lumi One Super Chemi Lumi One Ultra Product No 07880 Product No 02230 Product No 11644 Sensitivity picogram mid femtogram low femtogram Components Reagents Main composition Package Solution A Luminol solution brown plastic bottle Solution B Peroxide solution white plastic bottle Required reagents Washing buffer Use either a or b a 0 05 Tween 20 in TBS TBS T Mix 50ml of Tris Buffered Saline 10x PH 7 4 Product No 35438 0 25g of Polyoxyethylene Sorbitan Monolaurate Tween 20 and distilled water to a final volume of 500ml b 0 05 Tween 20 in PBS PBS T Mix 50ml of Phosphate Buffered Saline 10x PH 7 4 Product No 27575 0 25g of Polyoxyethylene Sorbitan Monolaurate Tween 20 and distilled water to a final volume of 500ml Blocking buffer Use one of the followings a Blocking One Product No 03953 b Blocking One P Product No 05999 for phosphoprotein detection c 2 5 skim milk in TBS T or PBS T d 2 5 BSA in
4. e desk and place the membrane protein side up on it Cover the membrane completely with the Working solution and incubate for 1 minute 11 Remove the Working solution from the membrane carefully with paper towels and envelop the membrane with a new plastic wrap Note Carefully remove all air bubbles and wrinkles from the enveloped membrane 12 Detection X ray film In a dark room place the enveloped membrane protein side up on a film cassette Place an X ray film on it and expose it for 3 minutes optimize the exposure time if necessary CCD imager Set the enveloped membrane protein side up in CCD imager and operate it according to the user manual CONAN KR WD o Attention Equilibrate the reagents to room temperature before preparing Working solutions Wear gloves and use tweezers when handling blotting membrane The dilution factor should be optimized for each antibody The following dilution ranges are recommended Primary antibody 1 1 000 1 5 000 Secondary antibody 1 20 000 1 100 000 If the signal is weak or too strong optimize the exposure time first If no improvement is observed optimize the concentration of antibodies and sample proteins Cautions Wear gloves and protective goggles because the product includes corrosive material Storage Refrigerator Protect from light Expiration One year from manufacturing Expiration date is stated on the product label Exp yy mm Packing KIT Product No 078
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