山羊F(ab')2抗小鼠IgM mu chain (HRP) (ab5930)

Thank you for your enquiry. Well done on getting your western blot to work. I have read your comments and have examined your blot images. I would like ot take each of your antibodies one at a time: ab3568 - Cytochrome P450 1A1 antibody (ab3568). You have obtained good signal using 20ug of lysate. The blot that you have e-mailed me would have fewer bands were the exposure to DAB have been reduced. We generally advise the use of chemiluminescence (ECL+). This is a more sensitive approach with which you can perform many exposures of the film to the blot. A shortened exposure would have generated significantly fewer bands. Given that you have detected bands on the blot very easily; to improve the specificity of the antibody and decrease the number of non-specific bands may I suggest that you use the antibody diluted to >1:500 e.g. 1:800 or 1:1000. This should serve to improve the recognition of the antibody for its target. I think that the reason why you are observing differences in the "length" of the bands and the number of cross reacting bands is due to the fact that the mass of the lysates that you are using are subtly different. The "longer" bands are due to diffusion of the lysates into adjacent areas and it is no consequence that there are more cross reacting bands on account of the larger mass. May I suggest that you try using the antibody diluted >1:500 and use an ECL or ECL+ detection system (provided you have a dark room), perform different exposure lengths and normalise your lysates so that you can be confident that they are all 20ug. To quantitate the mass of protein you will need to perform a Bradford assay or similar protein quant. With regards the telomerase, please can you tell me which of the antibodies that you were using e.g.abxxxx. I cannot imagine that the reason that you do not have any bands on the second blot is due to the use of BSA. BSA often gives a better, cleaner more specific blot. Can you tell me whether this blot gave a good Ponceau S staining? Were there any differences in the two blots. The transfer is the most likely problem, but if you tell me the antibody that you used I can check the datasheet to determine whether I can offer any more tips specific to the antibody e.g. dilution etc. I look forward to hearing from you with further details. I hope this information helps, please do not hesitate to contact me should you require further assistance.