Shorter-wavelength amine-reactive fluorophores are less frequently used for preparing bioconjugates because dyes excited with longer wavelengths, and therefore lower energy, are widely available and less likely to cause photodamage to labeled biomolecules. Moreover, many cells and tissues autofluoresce when excited with ultraviolet (UV) light, producing detection-confounding background signals. However, for certain multicolor fluorescence applications—including immunofluorescence, nucleic acid and protein microarrays, in situ hybridization and neuronal tracing—a blue-fluorescent probe provides a contrasting color that is clearly resolved from the green, yellow, orange or red fluorescence of the longer-wavelength probes.

The short-wavelength reactive dyes that we recommend for preparing the brightest blue-fluorescent bioconjugates are the Alexa Fluor 350, Alexa Fluor 405, AMCA-X, Marina Blue, Pacific Blue and Cascade Blue derivatives (Amine-reactive, ultraviolet light-excitable fluorophores for labeling proteins and nucleic acids—Table 1.12). Alexa Fluor 430, Pacific Orange and Cascade Yellow dyes fill a spectral void because they exhibit the rare combination of absorption between 400 nm and 450 nm and fluorescence emission beyond 500 nm. The amine-reactive naphthalene, pyrene and Dapoxyl derivatives are important for the production of environment-sensitive probes in protein structure and function studies (Amine-reactive, environment-sensitive fluorophores—Table 1.13); their thiol-reactive counterparts are discussed in Thiol-Reactive Probes Excited with Ultraviolet Light—Section 2.3. Many of our UV light–excitable reactive dyes are more commonly employed for such bioanalytical techniques as HPLC derivatization, amino acid sequencing and protein determination and are therefore discussed in Reagents for Analysis of Low Molecular Weight Amines—Section 1.8.

Alexa Fluor 350 and Other Coumarin Derivatives

Alexa Fluor 350 and AMCA-X Dyes

Derivatives of 7-aminocoumarin dyes are widely used labeling reagents for preparing protein and nucleic acid conjugates,ref and we offer two important amine-reactive 7-aminocoumarin derivatives: Alexa Fluor 350 carboxylic acid succinimidyl ester ref (A10168) and AMCA-X succinimidyl ester (AMCA-X, SE; A6118).

The sulfonated coumarin derivative, Alexa Fluor 350 carboxylic acid succinimidyl ester (structure), is more water soluble than either AMCA succinimidyl ester or AMCA-X succinimidyl ester (structure) and yields protein conjugates that are more fluorescent than those prepared from its nonsulfonated analog (Figure 1.7.1). Alexa Fluor 350 protein conjugates are optimally excited at 346 nm and have bright blue fluorescence emission (Figure 1.7.2, photo) at wavelengths slightly shorter than AMCA or AMCA-X conjugates (spectra) (442 nm versus 448 nm), which reduces the dye's spectral overlap with the emission of fluorescein. We offer several reactive versions of Alexa Fluor 350 dye, including:

AMCA-X succinimidyl ester (A6118) contains a seven-atom aminohexanoyl spacer ("X") between the fluorophore and the reactive group. This spacer separates the fluorophore from the biomolecule to which it is conjugated, potentially reducing the quenching that typically occurs upon conjugation and making the dye more available for recognition by secondary detection reagents. Slightly longer-wavelength conjugates can be prepared from the isothiocyanate (DACITC, D10166), succinimidyl esters (D374, D1412) or free acids (D126, D1421) of 7-dialkylaminocoumarins.ref

Alexa Fluor 350 streptavidin  


Figure 1.7.1
Comparison of the relative fluorescence of 7-amino-4-methylcoumarin-3-acetic acid (AMCA) streptavidin (open circle) and Alexa Fluor 350 streptavidin, a sulfonated AMCA derivative (S11249, filled circle). Conjugate fluorescence is determined by measuring the fluorescence quantum yield of the conjugated dye relative to that of the free dye and multiplying by the number of fluorophores per protein.


Figure 1.7.2 Absorption spectra of our ultraviolet and blue light–absorbing Alexa Fluor dyes.

Alexa Fluor 430 Dye

Few reactive dyes that absorb between 400 nm and 450 nm have appreciable fluorescence beyond 500 nm in aqueous solution. Alexa Fluor 430 dye fills this spectral gap.ref Excitation near its absorption maximum at 431 nm (Figure 1.7.2) is accompanied by strong green fluorescence with an emission maximum at 541 nm. The amine-reactive succinimidyl ester of Alexa Fluor 430 carboxylic acid (A10169, structure) is available, as well as Alexa Fluor 430 conjugates of secondary antibodies (A11063, A11064; Secondary Immunoreagents—Section 7.2, Summary of Molecular Probes secondary antibody conjugates—Table 7.1) and streptavidin (S11237, Avidin, Streptavidin, NeutrAvidin and CaptAvidin Biotin-Binding Proteins and Affinity Matrices—Section 7.6, Molecular Probes avidin, streptavidin, NeutrAvidin and CaptAvidin conjugates—Table 7.9).

Marina Blue and Pacific Blue Dyes

Marina Blue and Pacific Blue dyes, both of which are based on the 6,8-difluoro-7-hydroxycoumarin fluorophore, exhibit bright blue fluorescence emission near 460 nm ref (Amine-reactive, ultraviolet light-excitable fluorophores for labeling proteins and nucleic acids—Table 1.12). The Marina Blue dye is optimally detected using optical filters configured for DAPI, whereas the Pacific Blue dye is ideally suited for 405 nm violet diode laser excitation on the Applied Biosystems Attune acoustic focusing cytometer and similarly equipped fluorescence microscopes (Figure 1.7.7). Significantly, the pKa values of these 6,8-difluoro-7-hydroxycoumarin derivatives are 2–3 log units lower than those of the corresponding 7-hydroxycoumarins (Figure 1.7.3). Thus, the Marina Blue and Pacific Blue dyes yield conjugates that are strongly fluorescent, even at neutral pH. For preparing bioconjugates, we offer amine-reactive succinimidyl esters of the Marina Blue and Pacific Blue dyes (M10165, structure; P10163, structure).

pH-dependent fluorescence  


Figure 1.7.3 Comparison of the pH-dependent fluorescence responses of 7-hydroxy-4-methylcoumarin (H189) and 6,8-difluoro-7-hydroxy-4-methylcoumarin (DiFMU, D6566). Fluorescence intensities were measured for equal concentrations of the two dyes using excitation/emission at 355/460 nm. Compared with 7-hydroxy-4-methylcoumarin (pKa = 7.8), DiFMU (pKa = 4.9) provides superior detection sensitivity in the 5–8 pH range under these conditions.

Other Hydroxycoumarin and Alkoxycoumarin Derivatives

The hydroxycoumarins (H185, H1193, H1428) exhibit pH-sensitive spectral properties, but the methoxycoumarins (M1410, M1420MP) do not.ref Hydroxycoumarins are often used to prepare reactive intermediates for the synthesis of radioiodinated materials.ref The spectral properties of the hydroxycoumarins allow their quantitation prior to radioiodination.ref

Alexa Fluor 350, Alexa Fluor 430 and Pacific Blue Protein Labeling Kits

For easy and trouble-free labeling of proteins with succinimidyl esters of the Alexa Fluor 350, Alexa Fluor 430 and Pacific Blue dyes, we offer Alexa Fluor 350, Alexa Fluor 430 and Pacific Blue Protein Labeling Kits (A10170, A10171, P30012; Active esters and kits for labeling proteins and nucleic acids—Table 1.2). These kits, which are described in greater detail in Kits for Labeling Proteins and Nucleic Acids—Section 1.2, contain everything that is required to perform three separate labeling reactions and to purify the resulting conjugates (Molecular Probes kits for protein and nucleic acid labeling—Table 1.3). The Alexa Fluor 350 and Pacific Bue Antibody Labeling Kits (A20180, P30013) can be used to prepare blue-fluorescent conjugates of monoclonal and polyclonal antibodies, as well as of other proteins in limited quantities (five labeling reactions of ~100 µg each). The APEX Pacific Blue Antibody Labeling Kit (A10478) utilizes a solid-phase technique to label 10–20 µg IgG antibody, even in the presence of stabilizing proteins or amine-containing buffers.

The Zenon Alexa Fluor 350, Zenon Alexa Fluor 430 and Zenon Pacific Blue Antibody Labeling Kits (Zenon Antibody Labeling Kits—Table 7.7) permit the rapid and quantitative labeling of antibodies—even submicrogram amounts—using a purified antibody fraction or a crude antibody preparation such as serum, ascites fluid or a hybridoma supernatant. These kits, along with Zenon technology, are described in detail in Zenon Technology: Versatile Reagents for Immunolabeling—Section 7.3.

Pacific Orange Dye

The succinimidyl ester of the Pacific Orange dye (P30253) yields conjugates with excitation/emission maxima of ~400/551 nm, making it ideal for use with 405 nm violet diode laser–equipped flow cytometers ref and fluorescence microscopes. Moreover, Pacific Blue and Pacific Orange conjugates can be simultaneously excited at 405 nm and emit at 455 nm and 551 nm, respectively, facilitating two-color analysis.

Several of our kits facilitate protein labeling with the Pacific Orange succinimidyl ester, including the Pacific Orange Protein Labeling Kit (P30016), the Pacific Orange Antibody Labeling Kit (P30014) and the Zenon Antibody Labeling Kits (Zenon Antibody Labeling Kits—Table 7.7), all of which are described in greater detail in Kits for Labeling Proteins and Nucleic Acids—Section 1.2.

Cascade Blue and Other Pyrene Derivatives

Cascade Blue Acetyl Azide

Cascade Blue acetyl azide is the amine-reactive sulfonated pyrene derivative ref that we use to prepare  blue-fluorescent Cascade Blue dye–labeled proteins and dextrans. The polar nature of this reagent makes it difficult to purify to homogeneity; however, we offer a Cascade Blue acetyl azide preparation (C2284, Amine-reactive, ultraviolet light-excitable fluorophores for labeling proteins and nucleic acids—Table 1.12, structure) that is ~60% reactive and packaged according to the net weight of the reactive dye. The remaining constituents are inorganic salts or unreactive forms of the dye that can readily be removed following conjugation.

As compared with the aminocoumarin derivatives, the Cascade Blue fluorophore shows less spectral overlap with fluorescein (Figure 1.7.4), an important advantage for multicolor applications. In addition, this reactive Cascade Blue derivative has high absorptivity (spectra), is highly fluorescent and resists quenching upon protein conjugation (Figure 1.7.5). Even at low degrees of labeling, Cascade Blue conjugates are significantly more fluorescent than are those of 7-amino-4-methylcoumarin-3-acetic acid (AMCA),ref and they remain preferred reagents for multicolor flow cytometry.ref

Fluorescence emission spectra  


Figure 1.7.4 Normalized fluorescence emission spectra of Cascade Blue (CB), 7-amino-4-methylcoumarin (AMC) and fluorescein in aqueous solutions.

Fluorescence per fluorophore
Figure 1.7.5 Histograms showing the fluorescence per fluorophore for A) fluorescein and B) Cascade Blue conjugated to various proteins, relative to the fluorescence of the free dye in aqueous solution, represented by 100 on the y-axis. The proteins represented are: 1) avidin, 2) bovine serum albumin, 3) concanavalin A, 4) goat IgG, 5) ovalbumin, 6) protein A, 7) streptavidin and 8) wheat germ agglutinin.

Alexa Fluor 405 Dye

With excitation/emission maxima of 402/421 nm (Figure 1.7.2), Alexa Fluor 405 dye is well matched to the 405 nm spectral line of violet diode lasers for fluorescence microscopy and flow cytometry.ref Alexa Fluor 405 succinimidyl ester is an amine-reactive derivative of our Cascade Blue dye. Not only is it offered at higher purity than the alternative Cascade Blue acetyl azide, but Alexa Fluor 405 succinimidyl ester also contains a 4-piperidinecarboxylic acid spacer that separates the fluorophore from its reactive moiety (structure). This spacer enhances the reactivity of the succinimidyl ester and minimizes any interactions between the fluorophore and the biomolecule to which it is conjugated.

As with Cascade Blue acetyl azide, Alexa Fluor 405 dye shows minimal spectral overlap with green fluorophores, making it ideal for multicolor applications.ref However, the violet fluorescence of Cascade Blue and Alexa Fluor 405 dyes is less visible to the human eye in fluorescence microscopy applications than the blue fluorescence of Alexa Fluor 350 and AMCA-X dyes. Alexa Fluor 405 dye is available as:

We also prepare Alexa Fluor 405 conjugates of secondary antibodies (Secondary Immunoreagents—Section 7.2, Summary of Molecular Probes secondary antibody conjugates—Table 7.1) and streptavidin (Avidin, Streptavidin, NeutrAvidin and CaptAvidin Biotin-Binding Proteins and Affinity Matrices—Section 7.6, Molecular Probes avidin, streptavidin, NeutrAvidin and CaptAvidin conjugates—Table 7.9). Alexa Fluor 405 conjugates are recognized by the anti–Alexa Fluor 405/Cascade Blue dye antibody (A5760, Anti-Dye and Anti-Hapten Antibodies—Section 7.4).

Other Pyrenes

Conjugates of the pyrene succinimidyl esters (P130, P6114)  have exceptionally long excited-state lifetimes (sometimes >100 nanoseconds), relatively short-wavelength emission and capacity for proximity-dependent excimer formation (Figure 1.7.6). These amine-reactive pyrene derivatives have primarily been used for labeling and detecting oligonucleotides,ref biogenic amines ref and polyamines.ref Pyrene binds strongly to carbon nanotubes via pi-stacking interactions. This property makes 1-pyrenebutanoic acid succinimidyl ester (P130) a valuable reagent for functionalizing these remarkable nanomaterials for coupling to proteins.ref

The long fluorescence lifetime of pyrenebutyric acid (1-pyrenebutanoic acid) permits time-gating of the fluorescence, which is a useful technique for discriminating between the dye signal and sample autofluorescence,ref and has been exploited for fluorescence immunoassays.ref For preparing pyrene conjugates with long fluorescence lifetimes, we recommend the more water-soluble succinimidyl ester of N-(1-pyrenebutanoyl)cysteic acid (P6114, structure). The amine-reactive 1-pyrenesulfonyl chloride (P24, Reagents for Analysis of Low Molecular Weight Amines—Section 1.8) has been used to generate a fluorescent ATP sensor via modification of an ATP-binding ribonucleopeptide.ref

Pyrene in ethanol  


Figure 1.7.6 Excimer formation by pyrene in ethanol. Spectra are normalized to the 371.5 nm peak of the monomer. All spectra are essentially identical below 400 nm after normalization. Spectra are as follows: 1) 2 mM pyrene, purged with argon to remove oxygen; 2) 2 mM pyrene, air-equilibrated; 3) 0.5 mM pyrene (argon-purged); and 4) 2 µM pyrene (argon-purged). The monomer-to-excimer ratio (371.5 nm/470 nm) is dependent on both pyrene concentration and the excited-state lifetime, which is variable because of quenching by oxygen.

Cascade Yellow and Other Pyridyloxazole Derivatives

Cascade Yellow Dye

Like the Alexa Fluor 430 and Pacific Blue dyes described above, the Cascade Yellow dye exhibits an excitation maximum that falls between those of the UV light–excited dyes and the fluoresceins. This sulfonated pyridyloxazole (PyMPO) laser dye (structure) exhibits an absorption maximum near 410 nm and an unusually high Stokes shift, with relatively strong emission at 550–570 nm ref (spectra). The large Stokes shift permits detection at a wavelength well beyond that of most sample autofluorescence, and allows multiple fluorophores to be excited at the same wavelengths and detected at different wavelengths. For example, protein conjugates of Cascade Yellow succinimidyl ester (C10164) can be simultaneously excited at 405 nm with Pacific Blue conjugates, and then separately detected at longer wavelengths (Figure 1.7.7). Cascade Yellow and Cascade Blue antibody conjugates, along with several phycobiliprotein tandem conjugates, are utilized in an 11-color polychromatic flow cytometry technique.ref

Pacific Blue goat anti–mouse IgG antibody  
Figure 1.7.7 Normalized fluorescence emission spectra of Pacific Blue goat anti–mouse IgG antibody (P10993) and a Cascade Yellow goat anti–mouse IgG antibody conjugate prepared with the Cascade Yellow succinimidyl ester (C10164). Both fluorescent conjugates are excited at 405 nm. When samples containing equal concentrations of antibody are compared, the peak fluorescence intensity of the Pacific Blue conjugate at 456 nm is nine times greater than that of the Cascade Yellow conjugate at 548 nm.

PyMPO Dye

The pyridyloxazole derivatives—including the succinimidyl ester (PyMPO, SE; S6110; structure) and the thiol-reactive maleimide (M6026, Thiol-Reactive Probes Excited with Visible Light—Section 2.2)—fill the spectral gap between UV light–excited dyes and the fluoresceins. These derivatives of the laser dye PyMPO exhibit absorption maxima near 415 nm and unusually high Stokes shifts, with emission at 560–580 nm.ref Like the naphthalene-based dyes, the pyridyloxazole dyes exhibit environment-sensitive fluorescence spectra. PyMPO SE has been used to synthesize fluorescent gramicidin derivatives for following ion channel–gating processes.ref

Naphthalenes, Including Dansyl Chloride

Aminonaphthalene-based probes tend to have emission spectra that are sensitive to the environment and to exhibit weak fluorescence in aqueous solution. Spectra of environment-sensitive probes respond to perturbations in the local environment (Amine-reactive, environment-sensitive fluorophores—Table 1.13). For example, changes in solvation that occur because of ligand binding, protein assembly or protein denaturation can often evoke changes in the fluorescence properties of these probes. This property has made dansyl chloride (5-dimethylaminonaphthalene-1-sulfonyl chloride, D21) and other aminonaphthalene-based dyes important tools for protein structural studies.

Dansyl chloride is nonfluorescent until it reacts with amines. The resulting dansyl amides have environment-sensitive fluorescence quantum yields and emission maxima, along with large Stokes shifts. Despite the weak absorptivity (ε ~4000 cm-1M-1 at 330–340 nm) and moderate fluorescence quantum yield of dansyl sulfonamides, dansyl chloride is widely used as a derivatization reagent for end-group analysis of proteins, amino acid analysis and HPLC detection (Reagents for Analysis of Low Molecular Weight Amines—Section 1.8). The succinimidyl ester of dansylaminohexanoic acid (dansyl-X, SE; D6104; structure) contains a seven-atom spacer ("X") that places the dansyl fluorophore further from its reaction site, potentially reducing the interaction of the fluorophore with the biomolecule to which it is conjugated and enhancing accessibility to antibody binding.ref A rabbit polyclonal antibody to the 1,5-dansyl fluorophore (A6398) that significantly enhances the dye's fluorescence is described in Anti-Dye and Anti-Hapten Antibodies—Section 7.4.

Conjugates of two isomers of dansyl chloride (2,5-dansyl chloride, D22; 2,6-dansyl chloride, D23) have smaller Stokes shifts and appreciably longer fluorescence lifetimes (up to ~30 nanoseconds) than conjugates of 1,5-dansyl chloride, making these isomers among the best available probes for fluorescence depolarization studies.ref These dyes are particularly useful for preparing fluorescent drug or ligand analogs that are expected to bind to hydrophobic sites in proteins or membranes. The lipophilicity of these reagents may also facilitate the labeling of sites within the membrane-spanning portions of cellular proteins.

Dapoxyl Dye

Dapoxyl dye (structure) is a particularly versatile derivatization reagent and precursor to environment-sensitive probes.ref Like Cascade Yellow dye, Dapoxyl dye exhibits an exceptionally large Stokes shift, with excitation/emission maxima of ~370/580 nm (spectra). Sulfonamides from Dapoxyl sulfonyl chloride (D10160) have much higher extinction coefficients than those of dansyl chloride (~26,000 cm-1M-1 versus about 4000 cm-1M-1) and equal or greater quantum yields when dissolved in organic solvents; however, the fluorescence of Dapoxyl derivatives is very sensitive to the dye environment, and fluorescence in water is very low, making them useful for sensing conformational changes,ref denaturation and phosphorylation states ref of proteins.

In addition to Dapoxyl sulfonyl chloride, we offer the amine-reactive Dapoxyl succinimidyl ester (D10161) and the carboxylic acid–reactive Dapoxyl (2-aminoethyl)sulfonamide (D10460, Derivatization Reagents for Carboxylic Acids and Carboxamides—Section 3.4). We have also exploited the environment-sensitive fluorescence of the Dapoxyl dye (Figure 1.7.8) to develop a highly selective and photostable stain for the endoplasmic reticulum (ER-Tracker Blue-White DPX, E12353; Probes for the Endoplasmic Reticulum and Golgi Apparatus—Section 12.4).

Emission spectra of Dapoxyl  
Figure 1.7.8
Normalized fluorescence emission spectra of Dapoxyl (2-aminoethyl)sulfonamide (D10460) in 1) hexane, 2) chloroform, 3) acetone, 4) acetonitrile and 5) 1:1 acetonitrile:water.

Bimane Derivative

Bimane mercaptoacetic acid (carboxymethylthiobimane, B30250; structure) is a blue-fluorescent dye with excitation/emission maxima of ~380/458 nm. It is useful as a reference standard for the fluorogenic monobromobimane and monochlorobimane reagents (Thiol-Reactive Probes Excited with Ultraviolet Light—Section 2.3) because it is an analog of the thioether product of their reaction with glutathione and other thiols.

Data Table

Cat # Links MW Storage Soluble Abs EC Em Solvent Notes
A6118 icon 443.46 F,D,L DMF, DMSO 353 19,000 442 MeOH  
A10168 icon icon 410.35 F,D,L H2O, DMSO 346 19,000 445 pH 7 1
A10169 icon icon 701.75 F,D,L H2O, DMSO 430 15,000 545 pH 7 1
A30000 icon icon 1028.26 F,DD,L H2O, DMSO 400 35,000 424 pH 7 1, 2, 3
B30250 icon icon 282.31 F,D,L DMSO 380 5700 458 MeOH  
C2284 icon icon 607.42 F,D,LL H2O, MeOH 396 29,000 410 MeOH 2, 4
C10164 icon icon 563.54 F,D,L DMF, DMSO 409 24,000 558 MeOH 5
D21 icon icon 269.75 F,DD,L DMF, MeCN 372 3900 none CHCl3 6, 7
D22 icon 269.75 F,DD,L DMF, MeCN 403 2900 none MeOH 7, 8
D23 icon 269.75 F,DD,L DMF, MeCN 380 16,000 none CHCl3 7, 8
D126 icon 247.25 L pH >6, DMF 370 22,000 459 MeOH  
D374 icon 344.32 F,D,L DMF, MeCN 376 22,000 468 MeOH  
D1412 icon 358.35 F,D,L DMSO, MeCN 442 64,000 483 pH 9 9
D1421 icon 261.28 L pH >6, DMF 409 34,000 473 pH 9  
D6104 icon icon 461.53 F,D,L DMF, MeCN 335 4200 518 MeOH  
D10160 icon icon 362.83 F,DD,L DMF, MeCN 403 22,000 see Notes MeOH 7, 10
D10161 icon 405.41 F,D,L DMF, DMSO 395 20,000 601 MeOH 11
D10166 icon 260.31 F,DD,L DMF, MeCN 400 36,000 476 MeOH 12, 13
H185 icon 206.15 L pH >6, DMF 386 29,000 448 pH 10 14
H1193 icon 303.23 F,D,L DMF, MeCN 419 36,000 447 MeOH  
H1428 icon 234.21 L pH >6, DMF 360 19,000 455 pH 10  
M1410 icon 317.25 F,D,L DMF, MeCN 358 26,000 410 MeOH  
M1420MP icon 220.18 L pH >6, DMF 336 20,000 402 pH 9  
M10165 icon icon 367.26 F,D,L DMF, MeCN 362 19,000 459 pH 9  
P24 icon 300.76 F,DD,L DMF, MeCN 350 28,000 380 MeOH 7, 15
P130 icon 385.42 F,D,L DMF, DMSO 340 43,000 376 MeOH 16
P6114 icon 574.65 F,D,L H2O, DMSO 341 38,000 376 MeOH 1, 16
P10163 icon icon 339.21 F,D,L DMF, MeCN 416 46,000 451 pH 9  
P30253 icon ~750 F,D,L H2O, DMSO 404 25,000 553 MeOH 1
S6110 icon 564.39 F,D,L DMF, DMSO 415 26,000 570 MeOH 5
  1. This sulfonated succinimidyl ester derivative is water soluble and may be dissolved in buffer at ~pH 8 for reaction with amines. Long-term storage in water is NOT recommended due to hydrolysis.
  2. The Alexa Fluor 405 and Cascade Blue dyes have a second absorption peak at about 376 nm with EC ~80% of the 395–400 nm peak.
  3. A30100 is an alternative packaging of A30000 but is otherwise identical.
  4. Unstable in water. Use immediately.
  5. Fluorescence emission spectrum shifts to shorter wavelengths in nonpolar solvents.
  6. D21 butylamine derivative has Abs = 337 nm (EC = 5300 cm-1M-1), Em = 492 nm in CHCl3. Em and QY are highly solvent dependent: Em = 496 nm (QY = 0.45) in dioxane, 536 nm (QY = 0.28) in MeOH and 557 nm (QY = 0.03) in H2O.ref EC typically decreases upon conjugation to proteins (EC = 3400 cm-1M-1 at 340 nm).ref Fluorescence lifetimes (τ) of protein conjugates are typically 12–20 nanoseconds.ref
  7. Do NOT dissolve in DMSO.
  8. D22 butylamine derivative: Abs = 375 nm (EC = 3100 cm-1M-1), Em = 470 nm in MeOH. D23 butylamine derivative: Abs = 375 nm (EC = 13,000 cm-1M-1), Em = 419 nm in CHCl3.
  9. D1412 reaction product with 1-butylamine has Abs = 427 nm (EC = 48,000 cm-1M-1), Em = 478 nm in pH 9 buffer.
  10. D10160 fluorescence is very weak. Reaction product with butylamine has Abs = 373 nm (EC = 26,000 cm-1M-1), Em = 551 nm.
  11. D10161 butylamine derivative: Abs = 367 nm (EC = 25,000 cm-1M-1), Em = 574 nm in MeOH. QY of the derivative is approximately 15-fold higher than the unreacted reagent.
  12. Isothiocyanates are unstable in water and should not be stored in aqueous solution.
  13. D10166 butylamine derivative: Abs = 376 nm (EC = 25,000 cm-1M-1), Em = 469 nm in MeOH. QY of the derivative is approximately 6-fold higher than the unreacted reagent.
  14. H185 Abs = 339 nm (EC = 19,000 cm-1M-1), Em = 448 nm at pH 4.
  15. Spectra of the reaction product with butylamine.
  16. Pyrene derivatives exhibit structured spectra. The absorption maximum is usually about 340 nm with a subsidiary peak at about 325 nm. There are also strong absorption peaks below 300 nm. The emission maximum is usually about 376 nm with a subsidiary peak at 396 nm. Excimer emission at about 470 nm may be observed at high concentrations.

Ordering Information

Catalog # Name Size List Price (USD) Qty
A10168 Alexa Fluor® 350 NHS Ester (Succinimidyl Ester) 5 mg 214.00
A30000 Alexa Fluor® 405 NHS Ester (Succinimidyl Ester) 1 mg 292.00
A30100 Alexa Fluor® 405 NHS Ester (Succinimidyl Ester) 5 mg 1,058.00
A10169 Alexa Fluor® 430 NHS Ester (Succinimidyl Ester) 5 mg 214.00
A20180 Alexa Fluor® 350 Antibody Labeling Kit 1 kit 407.00
A10170 Alexa Fluor® 350 Protein Labeling Kit 1 kit 394.00
A10171 Alexa Fluor® 430 Protein Labeling Kit 1 kit 472.00
A6118 AMCA-X, SE (6-((7-Amino-4-Methylcoumarin-3-Acetyl)amino)Hexanoic Acid, Succinimidyl Ester) 10 mg 195.00
C2284 Cascade Blue® Acetyl Azide, Trisodium Salt 5 mg 263.00
D10161 Dapoxyl® Carboxylic Acid, Succinimidyl Ester 5 mg 128.00
D1421 7-Diethylaminocoumarin-3-Carboxylic Acid 100 mg 87.00
D1412 7-Diethylaminocoumarin-3-Carboxylic Acid, Succinimidyl Ester 25 mg 163.00
D374 DMACA, SE, 7-Dimethylaminocoumarin-4-Acetic Acid, Succinimidyl Ester 25 mg 243.00
D21 Dansyl Chloride (5-Dimethylaminonaphthalene-1-Sulfonyl Chloride) 1 g 89.00
D23 2-Dimethylaminonaphthalene-6-Sulfonyl Chloride 100 mg 253.00
H185 7-Hydroxycoumarin-3-Carboxylic Acid, reference standard 100 mg 178.00
M10165 Marina Blue® Succinimidyl Ester 5 mg 228.00
M1420MP 7-Methoxycoumarin-3-Carboxylic Acid 100 mg 104.00
M1410 7-Methoxycoumarin-3-Carboxylic Acid, Succinimidyl Ester 25 mg 140.00
P10163 Pacific Blue™ Succinimidyl Ester 5 mg 240.00
P30253 Pacific Orange™ Succinimidyl Ester, Triethylammonium Salt 1 mg 231.00
P130 1-Pyrenebutanoic Acid, Succinimidyl Ester 100 mg 227.00
S6110 PyMPO, SE, 1-(3-(Succinimidyloxycarbonyl)Benzyl)-4-(5-(4-Methoxyphenyl)Oxazol-2-yl)Pyridinium Bromide 5 mg 209.00
Z25000 Zenon® Alexa Fluor® 350 Mouse IgG1 Labeling Kit 1 kit 300.00
Z25100 Zenon® Alexa Fluor® 350 Mouse IgG2a Labeling Kit 1 kit 300.00
Z25300 Zenon® Alexa Fluor® 350 Rabbit IgG Labeling Kit 1 kit 300.00
Z25013 Zenon® Alexa Fluor® 405 Mouse IgG1 Labeling Kit 1 kit 311.00
Z25113 Zenon® Alexa Fluor® 405 Mouse IgG2a Labeling Kit 1 kit 335.00
Z25213 Zenon® Alexa Fluor® 405 Mouse IgG2b Labeling Kit 1 kit 365.00
Z25313 Zenon® Alexa Fluor® 405 Rabbit IgG Labeling Kit 1 kit 310.00
Z25041 Zenon® Pacific Blue™ Mouse IgG1 Labeling Kit 1 kit 314.00
A10478 APEX™ Pacific Blue™ Antibody Labeling Kit 1 kit 389.00
Z25256 Zenon® Pacific Orange™ Mouse IgG1 Labeling Kit 1 kit 314.00
Z25257 Zenon® Pacific Orange™ Mouse IgG2a Labeling Kit 1 kit 319.00
P30012 Pacific Blue™ Protein Labeling Kit 1 kit 420.00
P30013 Pacific Blue™ Antibody Labeling Kit 1 kit 420.00
P30014 Pacific Orange™ Antibody Labeling Kit 1 kit 435.00
P30016 Pacific Orange™ Protein Labeling Kit 1 kit 440.00
For Research Use Only. Not for use in diagnostic procedures.