Lissamine

Lissamine Green B l-[bis-(4,4 -dimethylaminophenyl)methyl]-2-hydroxynapbthalene-3,6-disulfonic acid sodium salt. Acid Green 50 [3087-16-9] M 576.6, m >200°(dec). Cl... 

Three types of reactive spectrally distinct fluorophores, namely lissamine rhoda-mine (LR) 80, 7-dimethylaminocoumarin (DMAC) 81, and bodipy-630 (BDPY) 82 dyes, prepared by coupling 3-azidopropylamine or propargylamine to commercially available amine-reactive dyes were evaluated for the use in selective dye-labeling of newly synthesised proteins in Rat-1 fibroblasts. 

Fig. 6.3. Chemical structures of rhodamine and some derivatives. TAMRA = N,N,TV,A-tetramethylrhodamine. Lissamine rhodamine = 3,5-disulfonyl-N,N, A,A-tetramethylrhodamine.

Smith, S. N. and Steer, R. P. (2001). The photophysics of Lissamine rhodamine-B sulphonyl chloride in aqueous solution implications for fluorescent protein-dye conjugates. J. Photochem. Photobiol. A Chem. 139, 151-156. 

Four forms of amine-reactive rhodamine probes are commonly available. Two of them are based on the tetramethyl derivatives of the fundamental rhodamine structure, one is based on the sulforhodamine B or Lissamine derivative, and the last is the sulforhodamine 101 or Texas Red-type of derivative. All of them react under alkaline conditions with primary amines in proteins and other molecules to form stable, highly fluorescent complexes. 

Remove unreacted NHS-rhodamine and reaction by-products by gel filtration or dialysis. Lissamine Rhodamine B Sulfonyl Chloride... 

The Lissamine form of rhodamine B consists of diethyl modifications on the two nitrogens of the upper rings of the basic rhodamine molecule as well as two sulfonate groups added at the... 

Figure 9.16 Lissamine rhodamine B sulfonyl chloride reacts with amine-containing molecules to produce stable sulfonamide bonds.

The spectral characteristics of protein conjugates made with Lissamine rhodamine B derivatives are of longer wavelength than those of tetramethylrhodamine—more toward the red region of the spectrum. In addition, modified proteins have better chemical stability and are somewhat easier to purify than those made from TRITC (discussed previously). Lissamine derivatives also make more photostable probes than the fluorescein derivatives (Section 1, this chapter). 

Lissamine rhodamine B sulfonyl chloride is relatively insoluble in water, but may be dissolved in DMF prior to the addition of a small aliquot to an aqueous reaction. Do not dissolve in DMSO, as sulfonyl chlorides will readily react with this solvent (Boyle, 1966). The compound has a maximal absorptivity at 556 nm with an extremely high extinction coefficient of up to 93,000M em-1 (in methanol) in highly purified form. Its emission maximum occurs at 576 nm, emitting red luminescence. 

Lissamine rhodamine B sulfonyl chloride has been used in numerous applications, including multiple-labeling techniques in microscopy (Wessendorf, 1990), for confocal microscopy... 

The following protocol is a general guide for labeling biological macromolecules with Lissamine rhodamine B sulfonyl chloride. Optimization of the fluorophore incorporation level (F/P ratio) may have to be done for specific labeling experiments. 

Dissolve Lissamine rhodamine B sulfonyl chloride (Invitrogen) in DMF at a concentration of l-2mg/ml. Protect from light and use immediately. 

The intense Texas Red fluorophore has a QY that is inherently higher than the tetrameth-ylrhodamine or Lissamine rhodamine B derivatives. Texas Red s luminescence is shifted maximally into the red region of the spectrum, and its emission peak only minimally overlaps with that of fluorescein. This makes Texas Red derivatives among the best choices of labels for use in double-staining techniques. 

Lissamine rhodamine B sulfonyl hydrazine is a hydrazide derivative of sulforhodamine B that can spontaneously react with aldehyde- or ketone-containing molecules to form a covalent,... 

Lissamine rhodamine B sulfonyl hydrazine can react with aldehyde groups to form hydrazone linkages. 

Lissamine rhodamine B sulfonyl hydrazine is soluble in DMF. The reagent may be dissolved in this solvent as a concentrated stock solution before adding a small aliquot to an aqueous reaction medium. The compound itself and all solutions made with it should be protected from light to avoid decomposition of its fluorescent properties. 

Modification with Lissamine Rhodamine B Sulfonyl Chloride... 

Figure 23.7 Avidin (or (strept)avidin) can be labeled with Lissamine rhodamine sulfonyl chloride to form a fluorescent probe.

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