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BATCH NUMBER -- NOT SPECIFIED -- ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM Non-specific staining SAMPLE mouse small intestine PRIMARY ANTIBODY ab26122 DETECTION METHOD ABC Vector POSITIVE AND NEGATIVE CONTROLS USED 1)no positive 2) whole rabbit serum of the same delution as a negative control. ANTIBODY STORAGE CONDITIONS +4C FIXATION OF SAMPLE Paraformaldehyde ANTIGEN RETRIEVAL Heat mediated technique (steamer) PERMEABILIZATION STEP no BLOCKING CONDITIONS deluted normal serum - 30 minutes SECONDARY ANTIBODY Vector ABC Elite Rabbit Kit - antirabbit biotinnilated 30 minutes, room temperature HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? delutions of ab ADDITIONAL NOTES Could you make clear situation with the reference for this ab26122? References for Islet 1 antibody - Neural Stem Cell Marker (ab26122) General / background references: Thaler JP et al. A postmitotic role for Isl-class LIM homeodomain proteins in the assignment of visceral spinal motor neuron identity. Neuron 41:337-50 (2004). PubMed: 14766174 I did not find any mentioning of this antibody. In this paper they produce antibody against Ist2 but not Ist1. And in the paper below monoclonal Isl1 was used. Pfaff, S.L., Mendelsohn, M., Stewart, C.L., Edlund, T., and Jessell, T.M. (1996). Requirement for LIM homeobox gene Isl1 in motor neuron generation reveals a motor neuron-dependent step in interneuron differentiation. Cell 84, 309–320.
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ANSWER: |
Thank you for taking the time to complete our questionaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody. I would like to offer some suggestions to help optimise the results. I would also appreciate if you can confirm some details of the procedure. Could you tell me what dilutions of the antibody you have tried and how long did you incubate the primary? We recommend starting at 1:50 or 1:100 and incubating overnight at 4C. Also could you tell me more about your fixation step and what percent normal serum did you use? We recommend using 10% normal serum for 2 hours at room temperature. You also may want to try incubating the secondary longer. The reference listed on the data sheet is a general reference about the target protein and its use in research. This antibody was not used in that reference. I am not familiar with the second reference you list, because it uses a monoclonal, and this antibody is a polyclonal. I hope this information is helpful. Should the suggestions not improve the results, please do not hesitate to contact me again with further details. |
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