Shop All Amine-Reactive Fluorophores, Biotins, Quantum Dots, & Other Labels

Alexa Fluor™ 790 NHS Ester (Succinimidyl Ester) (Invitrogen™)

Alexa Fluor® 790 is a bright and photostable near-IR dye that is spectrally similar to indocyanine green (ICG) and the IRDye® 800 dye. Used for stable signal generation in imaging and flow cytometry, Alexa Fluor® 790 dye is water soluble and pH-insensitive from pH 4 to pH 10. Fluorescence of this long-wavelength Alexa Fluor® dye is not visible to the human eye but is readily detected by most imaging systems. As the longest-wavelength Alexa Fluor® dye, the emission is well separated from commonly used far-red fluorophores such as Alexa Fluor® 647 dye or allophycocyanin (APC), facilitating multicolor analysis. This fluorophore is also expected to be useful for small animal in-vivo imaging (SAIVI) applications or for two-color western applications with the LI-COR® Odyssey® infrared imaging system. In addition to reactive dye formulations, we offer Alexa Fluor® 790 dye conjugated to a variety of antibodies, peptides, proteins, tracers, and amplification substrates optimized for cellular labeling and detection.The NHS ester (or succinimidyl ester) of Alexa Fluor® 790 is the most popular tool for conjugating this dye to a protein or antibody. NHS esters can be used to label to the primary amines (R-NH2) of proteins, amine-modified oligonucleotides, and other amine-containing molecules. The resulting Alexa Fluor® conjugate will exhibit brighter fluorescence and greater photostability than the conjugates of other spectrally similar fluorophores.

Detailed information about this AlexaFluor® NHS ester:

Fluorophore label: Alexa Fluor® 790 dye
Reactive group: NHS ester
Reactivity: Primary amines on proteins and ligands, amine-modified oligonucleotides
Ex/Em of the conjugate: 784/814 nm
Extinction coefficient: 260,000 cm-1M-1
Spectrally similar dyes: indocyanine green (ICG) and the IRDye® 800 dye
Molecular weight: ~1750

Typical Conjugation Reaction
You can conjugate amine-reactive reagents with virtually any protein or peptide (the provided protocol is optimized for IgG antibodies). You can scale the reaction for any amount of protein, but the concentration of the protein should be at least 2 mg/mL for optimal results. We recommend trying three different degrees of labeling, using three different molar ratios of the reactive reagent to protein.

The Alexa Fluor® NHS ester is typically dissolved in high-quality anhydrous dimethylformamide (DMF) or dimethylsulfoxide (DMSO) (D12345), and the reaction is carried out in 0.1–0.2 M sodium bicarbonate buffer, pH 8.3, at room temperature for 1 hour. Because the pKa of the terminal amine is lower than that of the lysine epsilon-amino group, you may achieve more selective labeling of the amine terminus using a buffer closer to neutral pH.

Conjugate Purification
Labeled antibodies are typically separated from free Alexa Fluor® dye using a gel filtration column, such as Sephadex™ G-25, BioGel® P-30, or equivalent. For much larger or smaller proteins, select a gel filtration media with an appropriate molecular weight cut-off or purify by dialysis. We offer several purification kits optimized for different quantities of antibody conjugate:
Antibody Conjugate Purification Kit for 0.5-1 mg (A33086)
Antibody Conjugate Purification Kit for 20-50 µg (A33087)
Antibody Conjugate Purification kit for 50-100 µg (A33088)

Learn More About Protein and Antibody Labeling
We offer a wide selection of Molecular Probes® antibody and protein labeling kits to fit your starting material and your experimental setup. See our Antibody Labeling kits or use our Labeling Chemistry Selection Tool for other choices. To learn more about our labeling kits, read Kits for Labeling Proteins and Nucleic Acids—Section 1.2 in The Molecular Probes® Handbook.

We’ll Make a Custom Conjugate for You
If you can’t find what you’re looking for in our online catalog, we’ll prepare a custom antibody or protein conjugate for you. Our custom conjugation service is efficient and confidential, and we stand by the quality of our work. We are ISO 9001:2000 certified.

EZ-Link™ Plus Activated Peroxidase (Thermo Scientific™)

Thermo Scientific Pierce Plus Activated Peroxidase is an amine-reactive form of horseradish peroxidase (HRP) that provides coupling efficiencies of greater than 95% with antibodies and other proteins. This product consists of one vial, containing 1 mg of lyophilized HRP and is sufficient to modify 1 mg of Immunoglobulin (IgG)

Features of Thermo Scientific Pierce Plus Activated Peroxidase:

Activated HRP – periodate-treated, aldehyde-activated horseradish peroxidase, ready for conjugation to antibodies and other proteins at sites of primary amines (e.g., lysines)
Permanent conjugation – reacts efficiently (95%) with primary amines to form covalent amide bonds upon treatment with sodium cyanoborohydride (included in kit)
High activity HRP – enzyme activity is 120 to 200 units/mg; lyophilized, activated enzyme is stable for at least 12 months at -20°C
Convenient quantities – each 1 mg-quantity of activated enzyme is sufficient for reaction with 1 mg of IgG to produce about 0.5 mL of conjugate
Customizable – vary the molar ratios, reaction buffer and pH, and other parameters to acheive conjugates with different levels of HRP incorporation and activity

Plus Activated Peroxidase is horseradish peroxidase (HRP) enzyme, whose native carbohydrates (sugars) have been gently oxidized with periodate to produce amine-reactive aldehyde groups. These carbonyls of Plus Activated Peroxidase spontaneously and efficiently crosslink with primary amines on antibody or other proteins. The method is more effective than other amine-reactive chemistries, such as glutaraldehyde coupling, which often causes polymerization and a greater degree of conjugate inactivation. The pre-activated enzyme eliminates the difficulties inherent in preparing and validating activated peroxidase from scratch. The kit provides the enzyme, accessory reagents and protocol to easily produce high-performance conjugates for Western blotting, ELISA and other detection techniques.

Related Products
EZ-Link™ Plus Activated Peroxidase Kit

Alexa Fluor™ 660 NHS Ester (Succinimidyl Ester) (Invitrogen™)

Alexa Fluor® 660 is a bright and photostable far-red dye with excitation ideally suited to the 633 or 647 nm laser line. Used for stable signal generation in imaging and flow cytometry, Alexa Fluor® 660 dye is water soluble and pH-insensitive from pH 4 to pH 10. Fluorescence of this long-wavelength Alexa Fluor® dye is not visible to the human eye but is readily detected by most imaging systems. In addition to reactive dye formulations, we offer Alexa Fluor® 660 dye conjugated to a variety of antibodies, peptides, proteins, tracers, and amplification substrates optimized for cellular labeling and detection.The NHS ester (or succinimidyl ester) of Alexa Fluor® 660 is the most popular tool for conjugating this dye to a protein or antibody. NHS esters can be used to label to the primary amines (R-NH2) of proteins, amine-modified oligonucleotides, and other amine-containing molecules. The resulting Alexa Fluor® conjugate will exhibit brighter fluorescence and greater photostability than the conjugates of other spectrally similar fluorophores.

Detailed information about this AlexaFluor® NHS ester:

Fluorophore label: Alexa Fluor® 660 dye
Reactive group: NHS ester
Reactivity: Primary amines on proteins and ligands, amine-modified oligonucleotides
Ex/Em of the conjugate: 668/698 nm
Extinction coefficient: 132,000 cm-1M-1
Spectrally similar dyes: Cy5®
Molecular weight: ~1100

Typical Conjugation Reaction
You can conjugate amine-reactive reagents with virtually any protein or peptide (the provided protocol is optimized for IgG antibodies). You can scale the reaction for any amount of protein, but the concentration of the protein should be at least 2 mg/mL for optimal results. We recommend trying three different degrees of labeling, using three different molar ratios of the reactive reagent to protein.

The Alexa Fluor® NHS ester is typically dissolved in high-quality anhydrous dimethylformamide (DMF) or dimethylsulfoxide (DMSO) (D12345), and the reaction is carried out in 0.1–0.2 M sodium bicarbonate buffer, pH 8.3, at room temperature for 1 hour. Because the pKa of the terminal amine is lower than that of the lysine epsilon-amino group, you may achieve more selective labeling of the amine terminus using a buffer closer to neutral pH.

Conjugate Purification
Labeled antibodies are typically separated from free Alexa Fluor® dye using a gel filtration column, such as Sephadex™ G-25, BioGel® P-30, or equivalent. For much larger or smaller proteins, select a gel filtration media with an appropriate molecular weight cut-off or purify by dialysis. We offer several purification kits optimized for different quantities of antibody conjugate:
Antibody Conjugate Purification Kit for 0.5-1 mg (A33086)
Antibody Conjugate Purification Kit for 20-50 µg (A33087)
Antibody Conjugate Purification kit for 50-100 µg (A33088)

Learn More About Protein and Antibody Labeling
We offer a wide selection of Molecular Probes® antibody and protein labeling kits to fit your starting material and your experimental setup. See our Antibody Labeling kits or use our Labeling Chemistry Selection Tool for other choices. To learn more about our labeling kits, read Kits for Labeling Proteins and Nucleic Acids—Section 1.2 in The Molecular Probes® Handbook.

We’ll Make a Custom Conjugate for You
If you can’t find what you’re looking for in our online catalog, we’ll prepare a custom antibody or protein conjugate for you. Our custom conjugation service is efficient and confidential, and we stand by the quality of our work. We are ISO 9001:2000 certified.

EZ-Link™ NHS-SS-Biotin (Thermo Scientific™)

Thermo Scientific EZ-Link NHS-SS-Biotin enables simple and efficient biotinylation of antibodies, proteins and other primary amine-containing molecules, as well as intracellular labeling.

Features of EZ-Link NHS-SS-Biotin:

Protein labeling—biotinylate antibodies or other proteins for detection or purification using streptavidin probes or resins
Membrane-permeable—can be used to label inside cells (intracellular)
Amine-reactive—reacts with primary amines (-NH2), such as the side-chain of lysines (K) or the amino-termini of polypeptides
Cleavable—disulfide bond in spacer arm allows the biotin label to be removed using reducing agents such as DTT; only a small sulfhydryl group remains attached to the molecule
Solubility—must be dissolved in DMSO or DMF before further dilution in aqueous buffers
Medium length—spacer arm (total length added to target) is 24.3 angstroms; it consists of the native biotin valeric acid group extended by a 7-atom chain

NHS-SS-Biotin is succinimidyl 2-(biotinamido)-ethyl-1,3' -dithiopropionate, a compound that enables simple and efficient biotinylation of antibodies, proteins and other primary amine-containing molecules. The extended spacer arm (24.3 angstroms) of this reagent reduces steric hindrance associated with binding to avidin or other biotin-binding proteins and incorporates a reducing agent-cleavable disulfide bond (-S-S-) within the spacer, providing further versatility to the reagent.

We manufacture biotin reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.

N-Hydroxysulfosuccinimide (NHS) esters of biotin are the most popular type of biotinylation reagent. NHS-activated biotins react efficiently with primary amino groups (-NH2) in alkaline buffers to form stable amide bonds. Proteins (e.g., antibodies) typically have several primary amines that are available as targets for labeling, including the side chain of lysine (K) residues and the N-terminus of each polypeptide.

Varieties of biotin NHS-ester reagents differ in length, solubility, cell permeability and cleavability. Non-sulfonated NHS-biotins are cell permeable but must be dissolved in organic solvent such as DMSO or DMF. Sulfo-NHS biotins (and those with pegylated spacers) are directly water soluble but not membrane permeable. Varieties containing disulfide bonds can be cleaved using reducing agents, enabling the biotin group to be disconnected from the labeled protein.

DNP-X, SE, 6-(2,4-Dinitrophenyl)aminohexanoic Acid, Succinimidyl Ester (Invitrogen™)

The amine-reactive DNP-X succinimidyl ester can be used bioconjugates that can be detected with anti-dinitrophenyl (anti-DNP) antibodies. This reactive hapten contains an additional seven-atom aminohexanoyl spacer ("X") between the fluorophore and the succinimidyl ester group. This spacer helps to separate increase accessibility to anti-DNP antibodies.

Malachite Green isothiocyanate (Invitrogen™)

Malachite green is a nonfluorescent photosensitizer that absorbs at long wavelengths (~630 nm). Its photosensitizing action can be targeted to particular cellular sites by using the amine-reactive malachite green isothiocyanate to conjugate this label to antibodies or other biomolecules.

EZ-Link™ Biocytin (Thermo Scientific™)

Thermo Scientific EZ-Link Biocytin is a simple variant of biotin that contains a primary amine in its valeric acid chain, which provides the essential backbone for construction of certain long-chain and trifunctional biotinylation reagents.

Features of EZ-Link Biocytin:

Biotinylation—label molecules and surfaces for assays or affinity purification methods involving avidin or streptavidin probes and resins
Amine-activated—primary amine can be crosslinked to proteins and material surfaces using EDC and other crosslinkers
Lysine derivative—functions as a biotin-tagged amino acid; potentially useful as a control or quenching reagent in biotin assay methods
Medium length—spacer arm (total length added to target) is 20.1 angstroms, representing a 7-atom extension of the native biotin valeric acid

Biocytin is ε-N-[d-biotinyl]-L-lysine, a compound formed by conjugation of the epsilon amine of lysine to the valeric acid side chain of biotin. It contains terminal carboxyl and amino groups, which provide functional handles for derivatization or conjugation to proteins, surfaces and other molecules. Carbodiimide (EDC) and NHS-ester crosslinker chemistries are most often utilized for covalent modifications involving biocytin. The compound is also useful as an amino acid control or biotin standard in assay methods involving streptavidin binding.

We manufacture biotin reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.

Amino-biotin compounds can be conjugated to functional groups of proteins and other molecules in a variety of ways. The most common method is to crosslink the terminal primary amine to carboxyl groups using . Carboxyl groups (-COOH) occur in aspartate or glutamate residues and the carboxy-terminus of polypeptides. When activated with EDC (Part No. 22980), carboxylates react with amino (—NH2) groups to form amide bonds.

Biocytin has both carboxyl and amino groups. Therefore, to prevent self-conjugation of biocytin, EDC-mediated reaction schemes with this compound are usually done in two steps, aided by Sulfo-NHS (Part No. 24510): (1) Activate a carboxylate molecule using EDC and Sulfo-NHS, followed by complete removal or inactivation of the EDC reagent; then (2) Add Biocytin to allow its primary amine to react with the Sulfo-NHS ester-activated carboxyl groups. See NHS-ester Chemistry. Subsequently, the remaining carboxyl group of biocytin could be conjugated to yet another molecule. For example, biocytin is the starting material for synthesis of Sulfo-SBED (Part No. 33033), a trifunctional reagent.

pHrodo™ iFL Green STP Ester (amine-reactive) (Invitrogen™)

The amine-reactive pH-sensitive pHrodo iFL Green STP Ester dye is an improved version of pHrodo Green dye optimized for the creation of bioconjugates to be used in the study of antibody internalization, endocytosis, and phagocytosis. pHrodo iFL Green dye dramatically increases fluorescence as the pH of its surroundings become more acidic. It is more soluble than the original pHrodo Green dye, making it more useful for the labeling of antibodies that may otherwise precipitate out of solution during conjugation.

• Use pH-sensitive pHrodo iFL Green STP Ester to make pH-sensitive bioconjugates of your choice
• Get faster, more accurate results than with any other endocytosis or phagocytosis assay—no need for wash steps or quenchers
• Multiplex with red fluorescent markers such as RFP, pHrodo iFL Red STP, and many others

The increase in fluorescence of pHrodo iFL Green dye as the pH changes from basic to acidic correlates with the acidification of intracellular vesicles, making it a valuable tool for the study of endocytosis or phagocytosis and their regulation by environmental factors, drugs, or pathogens. The spectral properties of pHrodo iFL Green dye make it useful for multi-color experiments. pHrodo iFL Green dye can be detected using most standard green-fluorescent filter sets and has been validated for use in a variety of applications, including flow cytometry, fluorescent microscopy, and high content screening (HCS). The lack of fluorescence of pHrodo iFL Green dye in a typical extracellular environment eliminates the need for wash steps or quencher dyes in the experimental workflow.

pHrodo iFL Green STP Ester is an amine-reactive dye that can be used to make pHrodo iFL Green STP bioconjugates in aqueous buffer. The STP ester will react with primary amines on a protein, cell, or virus to create a stable conjugate that can be used in live cell assays or stored for later use.

Alexa Fluor™ 680 NHS Ester (Succinimidyl Ester) (Invitrogen™)

Alexa Fluor® 680 is a bright and photostable near-IR dye that is spectrally similar to the Cy5.5 dye. Used for stable signal generation in imaging and flow cytometry, Alexa Fluor® 680 dye is water soluble and pH-insensitive from pH 4 to pH 10. Fluorescence of this long-wavelength Alexa Fluor® dye is not visible to the human eye but is readily detected by most imaging systems. In addition to reactive dye formulations, we offer Alexa Fluor® 680 dye conjugated to a variety of antibodies, peptides, proteins, tracers, and amplification substrates optimized for cellular labeling and detection (learn more).

The NHS ester (or succinimidyl ester) of Alexa Fluor® 680 is the most popular tool for conjugating this dye to a protein or antibody. NHS esters can be used to label to the primary amines (R-NH2) of proteins, amine-modified oligonucleotides, and other amine-containing molecules. The resulting Alexa Fluor® conjugate will exhibit brighter fluorescence and greater photostability than the conjugates of other spectrally similar fluorophores.

Detailed information about this AlexaFluor® NHS ester:

Fluorophore label: Alexa Fluor® 680 dye
Reactive group: NHS ester
Reactivity: Primary amines on proteins and ligands, amine-modified oligonucleotides
Ex/Em of the conjugate: 684/707 nm
Extinction coefficient: 183,000 cm-1M-1
Spectrally similar dyes: Cy5.5

Typical Conjugation Reaction
You can conjugate amine-reactive reagents with virtually any protein or peptide (the provided protocol is optimized for IgG antibodies). You can scale the reaction for any amount of protein, but the concentration of the protein should be at least 2 mg/mL for optimal results. We recommend trying three different degrees of labeling, using three different molar ratios of the reactive reagent to protein.

The Alexa Fluor® NHS ester is typically dissolved in high-quality anhydrous dimethylformamide (DMF) or dimethylsulfoxide (DMSO) (D12345), and the reaction is carried out in 0.1–0.2 M sodium bicarbonate buffer, pH 8.3, at room temperature for 1 hour. Because the pKa of the terminal amine is lower than that of the lysine epsilon-amino group, you may achieve more selective labeling of the amine terminus using a buffer closer to neutral pH.

Conjugate Purification
Labeled antibodies are typically separated from free Alexa Fluor® dye using a gel filtration column, such as Sephadex™ G-25, BioGel® P-30, or equivalent. For much larger or smaller proteins, select a gel filtration media with an appropriate molecular weight cut-off or purify by dialysis. We offer several purification kits optimized for different quantities of antibody conjugate:
Antibody Conjugate Purification Kit for 0.5-1 mg (A33086)
Antibody Conjugate Purification Kit for 20-50 µg (A33087)
Antibody Conjugate Purification kit for 50-100 µg (A33088)

Learn More About Protein and Antibody Labeling
We offer a wide selection of Molecular Probes® antibody and protein labeling kits to fit your starting material and your experimental setup. See our Antibody Labeling kits or use our Labeling Chemistry Selection Tool for other choices. To learn more about our labeling kits, read Kits for Labeling Proteins and Nucleic Acids—Section 1.2 in The Molecular Probes® Handbook.

We’ll Make a Custom Conjugate for You
If you can’t find what you’re looking for in our online catalog, we’ll prepare a custom antibody or protein conjugate for you. Our custom conjugation service is efficient and confidential, and we stand by the quality of our work. We are ISO 9001:2000 certified.

Oregon Green™ 488 Carboxylic Acid, Succinimidyl Ester, 5-isomer (Invitrogen™)

This single 5-isomer preparation of our proprietary, amine-reactive dye, Oregon Green 488 carboxylic acid, succinimidyl ester and its conjugates have green fluorescence similar to that of fluorescein but are more photostable. Additionally, Oregon Green 488 dye has a lower pKa than fluorescein (pKa = 4.7 versus 6.4 for fluorescein), making this bright dye less pH sensitive in the physiological range.

EZ-Link™ NHS-SS-PEG4-Biotin (Thermo Scientific™)

Thermo Scientific EZ-Link NHS-SS-PEG4-Biotin includes disulfide and polyethylene glycol (PEG) groups in its spacer arm to provide biotinylation that is cleavable and which enhances solubility.

Features of EZ-Link NHS-SS-PEG4-Biotin:

Protein labeling—biotinylate antibodies or other proteins for detection or purification using streptavidin probes or resins
Amine-reactive—reacts with primary amines (-NH2), such as the side-chain of lysines (K) or the amino-termini of polypeptides
Cleavable—disulfide bond in spacer arm allows the biotin label to be removed using reducing agents such as DTT; only a small sulfhydryl group remains attached to the molecule
Pegylated—spacer arm contains a hydrophilic, 4-unit, polyethylene glycol (PEG) group
Enhances solubility—pegylation imparts water solubility to the biotinylated molecule, helping to prevent aggregation of biotinylated antibodies stored in solution
Long reach—spacer arm (total length added to target) is 37.9 angstroms; this reduces steric hindrance when binding to avidin molecules

NHS-SS-PEG4-Biotin enables simple and efficient biotin labeling of antibodies, proteins and other primary amine-containing macromolecules. The N-hydroxysuccinimide ester (NHS ester) group reacts specifically and efficiently with protein and peptide lysine (K) and N-terminal amino groups at pH 7-9 to form stable amide bonds. The hydrophilic polyethylene glycol (PEG) spacer arm imparts water solubility that is transferred to the biotinylated molecule, thus reducing aggregation of labeled proteins stored in solution. The integral disulfide bond enables the biotin group to be cleaved from the labeled molecule using DTT or other reducing agents, providing for efficient recovery of avidin-bound protein complexes in affinity-based assays.

We manufacture biotin reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.

N-Hydroxysulfosuccinimide (NHS) esters of biotin are the most popular type of biotinylation reagent. NHS-activated biotins react efficiently with primary amino groups (-NH2) in alkaline buffers to form stable amide bonds. Proteins (e.g., antibodies) typically have several primary amines that are available as targets for labeling, including the side chain of lysine (K) residues and the N-terminus of each polypeptide.

Varieties of biotin NHS-ester reagents differ in length, solubility, cell permeability and cleavability. Non-sulfonated NHS-biotins are cell permeable but must be dissolved in organic solvent such as DMSO or DMF. Sulfo-NHS biotins (and those with pegylated spacers) are directly water soluble but not membrane permeable. Varieties containing disulfide bonds can be cleaved using reducing agents, enabling the biotin group to be disconnected from the labeled protein.

Acryloyl-X, SE, 6-((acryloyl)amino)hexanoic Acid, Succinimidyl Ester (Invitrogen™)

The succinimidyl ester of acryloyl-X, SE (6-((acryloyl)amino)hexanoic acid) reacts with amines, of proteins, amine-modified nucleic acids and other biomolecules to yield acrylamides that can be copolymerized into polyacrylamide matrices or on surfaces, such as in microarrays and in biosensors.

LanthaScreen™ Thiol Reactive Tb Chelate

As part of the LanthaScreen® TR-FRET toolbox of assay reagents, LanthaScreen® Thiol Reactive Tb Chelate labeling reagent is available for assay development. The thiol-reactive maleimide group can be conjugated to virtually any peptide or protein that contains one or more accessible thiol moieties, such as the thiol group found on a cysteine residue. Energy transfer from the terbium donor to a suitable acceptor such as fluorescein is readily detected by monitoring an increase in acceptor fluorescence intensity. See the user guide for an application-based protocol for cysteine-containing peptides or proteins.

BODIPY™ FL-C5 NHS Ester (Succinimidyl Ester) (Invitrogen™)

BODIPY® FL-C5 dye is bright, green fluorescent dye with similar excitation and emission to fluorescein (FITC) or Alexa Fluor® 488 dye. It has a high extinction coefficient and fluorescence quantum yield and is relatively insensitive to solvent polarity and pH change. In contrast to the highly water soluble fluorophores Alexa Fluor® 488 dye and fluorescein (FITC), BODIPY® dyes have unique hydrophobic properties ideal for staining lipids, membranes, and other lipophilic compounds. BODIPY® FL-C5 dye has a relatively long excited-state lifetime (typically 5 nanoseconds or longer), which is useful for fluorescence polarization-based assays and a large two-photon cross-section for multiphoton excitation. In addition to reactive dye formulations, we offer BODIPY® FL-C5 dye conjugated to a variety of antibodies, peptides, proteins, tracers, and amplification substrates optimized for cellular labeling and detection (learn more).

The NHS ester (or succinimidyl ester) of BODIPY® FL-C5 is the most popular tool for conjugating the dye to a protein or antibody. NHS esters can be used to label the primary amines (R-NH2) of proteins, amine-modified oligonucleotides, and other amine-containing molecules. The resulting BODIPY® FL-C5 conjugates exhibit bright fluorescence, narrow emission bandwidths, and relatively long excited-state lifetimes, which can be useful for fluorescence polarization assays and two-photon excitation (TPE) microscopy.

This reactive dye contains a C5 alkyl spacer between the fluorophore and the NHS ester group. This spacer helps to separate the fluorophore from its point of attachment, potentially reducing the interaction of the fluorophore with the biomolecule to which it is conjugated.

Detailed information about this BODIPY® FL-C5 NHS ester:

Fluorophore label: BODIPY® FL-C5 dye
Reactive group: NHS ester (succinimidyl ester)
Reactivity: Primary amines on proteins and ligands, amine-modified oligonucleotides
Ex/Em of the conjugate: 504/511 nm
Extinction coefficient: 87,000 cm-1M-2
Molecular weight: 417.22

Typical Conjugation Reaction
Amine-reactive reagents can be conjugated with virtually any protein or peptide; the provided protocol is optimized for IgG antibodies. The reaction can be scaled for any amount of protein, but the concentration of the protein should be at least 2 mg/mL for optimal results. We recommend trying three different degrees of labeling, using three different molar ratios of the reactive reagent to protein.

The BODIPY® NHS ester is typically dissolved in high-quality anhydrous dimethylformamide (DMF) or dimethylsulfoxide (DMSO), and the reaction is carried out in 0.1-0.2 M sodium bicarbonate buffer, pH 8.3, at room temperature for 1 hour. Because the pKa of the terminal amine is lower than that of the lysine epsilon-amino group, you may achieve more selective labeling of the amine terminus using a buffer closer to neutral pH.

Conjugate Purification
Labeled antibodies are typically separated from free BODIPY® dye using a gel filtration column, such as Sephadex™ G-25, BioGel® P-30, or equivalent. For much larger or smaller proteins, select a gel filtration medium with an appropriate molecular weight cut-off or purify by dialysis. We offer several purification kits optimized for different quantities of antibody conjugate:
Antibody Conjugate Purification Kit for 0.5-1 mg (A33086)
Antibody Conjugate Purification Kit for 20-50 µg (A33087)
Antibody Conjugate Purification kit for 50-100 µg (A33088)

Learn More About Protein and Antibody Labeling
We offer a wide selection of Molecular Probes® antibody and protein labeling kits to fit your starting material and your experimental setup. See our Antibody Labeling kits or use our Labeling Chemistry Selection Tool for other choices. To learn more about our labeling kits, read Kits for Labeling Proteins and Nucleic Acids—Section 1.2 in The Molecular Probes® Handbook.

We’ll Make a Custom Conjugate for You
If you can’t find what you’re looking for in our online catalog, we’ll prepare a custom antibody or protein conjugate for you. Our custom conjugation service is efficient and confidential, and we stand by the quality of our work. We are ISO 9001:2000 certified.

Qdot™ 705 ITK™ Amino (PEG) Quantum Dots (Invitrogen™)

Qdot® 705 ITK™ amino (PEG) quantum dots are the ideal starting material for preparing custom conjugates of ultrabright and photostable fluorescently labeled proteins or other biopolymers. These probes are functionalized with amine-derivatized PEG, which prevents non-specific interactions and provides a convenient handle for conjugation. The amino quantum dots react efficiently with isothiocyanates and succinimidyl esters, or with native carboxylic acids using water-soluble carbodiimides such as EDC. Such derivatives may be used for various labeling and tracking applications that require ultrabright and stable fluorescence. Our Qdot® ITK™ amino quantum dots are provided as 8 µM solutions and are available in 8 colors of Qdot® probes.

Important Features of Qdot® ITK™ Amino Quantum Dots:
• Qdot® 705 ITK™ amino quantum dot has emission maxima of ~705 nm
• Extremely photostable and bright fluorescence
• Efficiently excited with single-line excitation sources
• Narrow emission, large stokes shift
• Available in multiple colors
• Ideal for various labeling and tracking applications


Properties of Qdot® Nanocrystals
Qdot® probes are ideal for imaging and labeling applications that require bright fluorescent signals and/or real-time tracking. Unique among fluorescent reagents, all nine available colors of Qdot® probes can be simultaneously excited with a single (UV to blue-green) light source. This property makes these reagents excellent for economical and user-friendly multiplexing applications. Qdot® labels are based on semiconductor nanotechnology and are similar in scale to moderately sized proteins.

About the Innovator’s Tool Kit Qdot® ITK™ Reagents
These Qdot® ITK™ probes are ideal for researchers who wish to prepare specific (non-stocked) conjugates for their applications and need customizable conjugation functionality.

Other Forms of Qdot® Nanocrystals are Available
In addition to the amine-derivatized form, we offer Qdot® ITK™ quantum dots with carboxyl and aliphatic hydrocarbon modifications. We’ve also developed a wide range of Qdot® nanocrystals conjugates and labeling kits. Investigate the properties of Qdot® nanocrystals or read the Molecular Probes® Handbook Section 6.6—Qdot® Nanocrystals to find out more.

For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.