| Technical Information on Probes Conjugated to Our Antibodies and Other Proteins: Tetramethyl Rhodamine Isothiocyanate (TRITC), Rhodamine Red-X (RRX), and Texas Red (TR) |
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| Conjugates of these rhodamine derivatives have different excitation (550, 570, and 596 nm) and emission (570, 590, and 620 nm) maxima (Table 1 and Figure 2). Although TRITC has been used traditionally with FITC for double labeling, better color separation is achieved by using RRX or Texas Red. However, it has been reported that use of Texas Red may lead to higher background staining (Wessendorf and Brelje, Histochemistry. 1992. 98, 81). We now recommend DyLight 594, instead of Texas Red, because it is brighter, more photostable, and more hydophilic than Texas Red. Table 1. Approximate peak wavelengths of excitation and emission for JIR conventional and DyLight fluorophores conjugated to affinity-purified antibodies. Only approximate values are given for purposes of comparing one fluorophore with another. Actual values may vary depending on the spectrofluorometer used in each laboratory. |
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| Rhodamine Red-X is particularly useful for three-color labeling with DyLight 488 and DyLight 649 by using a confocal microscope equipped with a krypton/argon laser. Fluorescence from RRX lies about midway between that of DyLight 488 and DyLight 649, and it shows little overlap with either dye (Figure 2). The krypton-argon laser emits lines at 488 nm, 568 nm, and 647 nm, all of which are optimal for exciting DyLight 488, RRX, and DyLight 649, respectively. By adding a 405 nm laser four-color labeling is possible using DyLight 405-conjugated secondary antibodies from JIR (Figure 2). | |||||||||||||||||||||||||||||||||||||||||||
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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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