BATCH NUMBER -- NOT SPECIFIED -- ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM Non-specific band SAMPLE Brain Cortex Homogenated PRIMARY ANTIBODY - DETECTION METHOD ECL POSITIVE AND NEGATIVE CONTROLS USED - ANTIBODY STORAGE CONDITIONS -20ºC SAMPLE PREPARATION Protease Inhibitors AMOUNT OF PROTEIN LOADED 2 mg/ml, 14microL each band. ELECTROPHORESIS/GEL CONDITIONS Gel acrylamide 10% TRANSFER AND BLOCKING CONDITIONS - SECONDARY ANTIBODY - HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 6 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? - ADDITIONAL NOTES -
Asked on Apr 12 2007
Thank you for your enquiry. I am sorry you are experiencing problems with this antibody. Often it is possible to make suggestions to improve your results. If we determine that the antibody is not working as stated on the datasheet, and you have purchased it in the last 90 days, we would be happy to replace or refund the antibody for you. In order for me to help you further, can you please confirm the followings: 1) how was the sample prepared? was SDS and beta-mercaptoethenol used? did you boiled it and for how long? what lysis buffer did you used? did you add protease inhibitor? 2) can you confirm that you used 28micro grams of protein per lane? 3) what secondary antibody did you used, as there was no information in the questionnaire? was it a anti-rabbit antibody? 4) how long did you incubate the primary and secondary antibodies? what was the dilution and temperature? We recommend incubating the primary overnight, at 4oC. 5) did you perform blocking procedures? what was the duration, blocking buffer, and concentration used?We recommend using 5% BSA in TBST for 1 hr. Can you also please submit an image of the blot showing the non-specific bands? I am looking forward to receiving the answers to the above questions and also an image of the non-specific bands so that we can resolve the matter. Have a nice day.
Answered on Apr 12 2007