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1. more bands 2. rat liver whole cell lysates 3. 50 micro grams 4. i used three different protease inhibitors and used sonication 5. no i didnt heat the samples 6. primary antibody raised in rabbit 7. 1 in 2000 8. incubated over night. did five to six washes 9. the secondary antibody concentrations were tested from 1 in 2000 to 1 in 10,000. 10. no it is not due to the secondary antibody as they look were no bands in secondary antibody control 11. incubated for one hour in secondary antibody 12. we are using horse raddish peroxidase and taking the pictures in x ray film 13. yes i did a column as control in which i didnt add the primary antibody 14. i tried different concentrations of secondary
Asked on Jul 14 2004
Thank you very much for the details that you have provided and for your patience. What was the lot number that you received? Why did you not heat the samples to denature the proteins? In addition to the non-specific bands that you are seeing, did you see the expected band at 46 kDa? I can offer you a replacement vial or a refund, please send me the Abcam order number of purchase order number that was used.
Answered on Jul 21 2004