| Multiple Labeling (ML) Using Labeled Secondary Antibodies |
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Simultaneous detection of more than one antigen depends on at least two important criteria:
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Note: Wash thoroughly after each step, including after the first blocking step. Do not dilute any antibody with normal serum. In this example, the secondary antibodies used in Steps 3, 6, and 9 do not recognize each other since they are all made in donkey. They have been solid-phase adsorbed so that they do not recognize the other primary antibodies used in Steps 2, 5, and 8. Also, they do not react with endogenous mouse Ig, which may be present in the mouse tissue. For a review of multi-color immunofluorescence labeling with confocal microscopy see Brelje, Wessendorf, and Sorenson, “Multi-color laser scanning confocal immunofluorescence microscopy: Practical application and limitations.” In Cell Biological Applications of Confocal Microscopy (Methods in Cell Biology. vol. 38). Ed. B. Matsumoto. Orlando, FL: Academic Press, Inc. 1993. , pp. 98-181. |
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DyLight™ |
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NEW Fluorescent Dyes |
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A new family of fluorescent dyes with improved brightness and photostability conjugated to Secondary Antibodies with recognized highest quality and diversity from Jackson ImmunoResearch Laboratories, Inc |
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