Name
BODIPY 530 Conjugation Kit (Fast ) (MAK-0247)
Cat
MAK-0247
Target
Antibody; Protein
Ex/Em (nm)
534/551
Product Type
Labeling Kit
Features
BODIPY 530 Conjugation Kit (Fast ) (MAK-0247) offers several standout features:
-Rapid Conjugation: achieve BODIPY 530 labeling in under 20 minutes with just 30 seconds of hands-on time.
-High Efficiency: ensures 100% antibody recovery, meaning no loss of valuable antibodies.
-Versatility: suitable for conjugating antibodies, proteins, and peptides. BODIPY 530 labeled antibodies can be used immediately in applications such as Western blot (WB), ELISA, and Immunohistochemistry (IHC) without further purification.
-Rapid Conjugation: achieve BODIPY 530 labeling in under 20 minutes with just 30 seconds of hands-on time.
-High Efficiency: ensures 100% antibody recovery, meaning no loss of valuable antibodies.
-Versatility: suitable for conjugating antibodies, proteins, and peptides. BODIPY 530 labeled antibodies can be used immediately in applications such as Western blot (WB), ELISA, and Immunohistochemistry (IHC) without further purification.
Background
BODIPY 530/550 dye is bright, orange fluorescent 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 530/550 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 530/550 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 BODIPY 530/550 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 530/550 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 C3 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.
Product Information
The Conjugation Kit (Fast) uses a simple and quick process for Fluorescein labeling / conjugation of antibodies. It can also be used to conjugate other proteins or peptides. To conjugate an antibody to fluorescein using this kit:
- add modifier to antibody and incubate for 60 mins
- Purify the conjugation product through a desalting column
The conjugated antibody can be used immediately in WB, ELISA, IHC, FACS, IF etc. No further purification is required and 100% of the antibody is recovered for use. Use the to learn more about the technology, or about conjugating other proteins and peptides to Fluorescein. Custom size conjugation kits up to 100 mg are available on demand. Please contact us to discuss your requirements.
- add modifier to antibody and incubate for 60 mins
- Purify the conjugation product through a desalting column
The conjugated antibody can be used immediately in WB, ELISA, IHC, FACS, IF etc. No further purification is required and 100% of the antibody is recovered for use. Use the to learn more about the technology, or about conjugating other proteins and peptides to Fluorescein. Custom size conjugation kits up to 100 mg are available on demand. Please contact us to discuss your requirements.
Size
Size: 1mg
Preactivated Fluorescein: 1 vial (Protect from light)
Buffer: 50 ul
DMSO: 150 ul
Purification Column+Tube: 2 vial
Preactivated Fluorescein: 1 vial (Protect from light)
Buffer: 50 ul
DMSO: 150 ul
Purification Column+Tube: 2 vial
Recommended Ratio
1. Using the maximum amount of antibody may result in less labelling per antibody.
2. Ideal antibody concentration is 1mg/ml. 0.5 - 1 mg/ml can be used if the maximum antibody volume is not exceeded. Antibodies > 2mg/ml or < 0.5 mg/ml should be diluted /concentrated.
2. Ideal antibody concentration is 1mg/ml. 0.5 - 1 mg/ml can be used if the maximum antibody volume is not exceeded. Antibodies > 2mg/ml or < 0.5 mg/ml should be diluted /concentrated.
Buffer Req.
Buffer should be pH 6.5-8.5.
Sample buffer
If a concentration is shown, then the constituent should be no more than the concentration shown. If several constituents are close to the limit of acceptable concentration, then this can inhibit conjugation.
(1) Compatible buffer constituents: 50mM / 0.6% Tris(Tris buffered saline is almost always ≤ 50 mM / 0.6%), 0.1% BSA, 50% glycerol, 0.1% sodium azide ( Sodium azide irreversibly inhibits HRP; antibodies with azide should be purified before using the HRP conjugation kit.), PBS.
(2) Potassium phosphate, Sodium chloride, HEPES, Sucrose, Sodium citrate, EDTA, Trehalose.
(3) Incompatible buffer constituents: Thiomerosal, Proclin, Glycine, Arginine, Glutathione, DTT.
(4) If a constituent of the buffer containing your antibody or protein is not listed above, please contact us.
(5) Only purified antibodies are suitable for use, ie. where other proteins, peptides, or amino acids are not present: antibodies in ascites fluid, serum or hybridoma culture media are incompatible.
(1) Compatible buffer constituents: 50mM / 0.6% Tris(Tris buffered saline is almost always ≤ 50 mM / 0.6%), 0.1% BSA, 50% glycerol, 0.1% sodium azide ( Sodium azide irreversibly inhibits HRP; antibodies with azide should be purified before using the HRP conjugation kit.), PBS.
(2) Potassium phosphate, Sodium chloride, HEPES, Sucrose, Sodium citrate, EDTA, Trehalose.
(3) Incompatible buffer constituents: Thiomerosal, Proclin, Glycine, Arginine, Glutathione, DTT.
(4) If a constituent of the buffer containing your antibody or protein is not listed above, please contact us.
(5) Only purified antibodies are suitable for use, ie. where other proteins, peptides, or amino acids are not present: antibodies in ascites fluid, serum or hybridoma culture media are incompatible.
Storage
Kits are intentionally shipped at ambient temperature with silica gel to avoid exposure to moisture. Upon receipt, store the kit frozen and protect from moisture. Before opening the outer container, allow the lyophilized components to reach room temperature to minimize condensation. -20°C
Procedure
(1) Take out the FLUORESCEIN and centrifuge for a few seconds to allow the dry powder of FLUORESCEIN in the tube to settle to the bottom;
(2) Add 100 ul of DMSO to the tube and mix repeatedly with a pipette or vortex until the FLUORESCEIN is completely dissolved;
(3) Add an appropriate amount of adjustment buffer to the antibody (10 ul of adjustment buffer per 100 ul of antibody);
(4) Add the dissolved FLUORESCEIN to the antibody (2.0-4.0 ul of FLUORESCEIN per 100 ug of antibody) and mix repeatedly with a pipette or vortex.
(5) Place the antibody-FLUORESCEIN mixture on a horizontal shaker or rotary mixer, and incubate it in the dark at room temperature (the reaction tube can be wrapped in aluminum foil) for 1 hour while shaking. Note: If a higher F/P ratio is required, the coupling time between antibody and FLUORESCEIN can be appropriately extended.
(6) Remove the purification column and centrifuge it at 3000 rpm for 2 minutes.
(7) Temporarily retain the buffer solution in the collection tube.
(8) Transfer the purification column to a new collection tube and aspirate the antibody-FLUORESCEIN conjugate onto the surface of the packing material in the purification column. If the sample volume is less than 50ul, first use 3.2 of the reserved buffer to make up the liquid volume to 50ul. The maximum sample volume should not exceed 100ul.
(9) After the sample has penetrated into the filler, centrifuge at 3000rpm for 2 minutes.
(10) Store the antibody-FLUORESCEIN conjugate in the dark at 4℃ for future use. The final product can be stored in the dark at 4℃ for up to 1 year.
(2) Add 100 ul of DMSO to the tube and mix repeatedly with a pipette or vortex until the FLUORESCEIN is completely dissolved;
(3) Add an appropriate amount of adjustment buffer to the antibody (10 ul of adjustment buffer per 100 ul of antibody);
(4) Add the dissolved FLUORESCEIN to the antibody (2.0-4.0 ul of FLUORESCEIN per 100 ug of antibody) and mix repeatedly with a pipette or vortex.
(5) Place the antibody-FLUORESCEIN mixture on a horizontal shaker or rotary mixer, and incubate it in the dark at room temperature (the reaction tube can be wrapped in aluminum foil) for 1 hour while shaking. Note: If a higher F/P ratio is required, the coupling time between antibody and FLUORESCEIN can be appropriately extended.
(6) Remove the purification column and centrifuge it at 3000 rpm for 2 minutes.
(7) Temporarily retain the buffer solution in the collection tube.
(8) Transfer the purification column to a new collection tube and aspirate the antibody-FLUORESCEIN conjugate onto the surface of the packing material in the purification column. If the sample volume is less than 50ul, first use 3.2 of the reserved buffer to make up the liquid volume to 50ul. The maximum sample volume should not exceed 100ul.
(9) After the sample has penetrated into the filler, centrifuge at 3000rpm for 2 minutes.
(10) Store the antibody-FLUORESCEIN conjugate in the dark at 4℃ for future use. The final product can be stored in the dark at 4℃ for up to 1 year.
Shipped
Ambient - Cannot Ship with Ice
Coupled Product Storage
Generally, conjugated antibodies can be stored at 4°C for up to 18 months. If longer storage is required, a cryoprotectant, such as 50% glycerol, can be used to store the conjugated antibody at -20°C. The optimal storage conditions for conjugated antibodies must be determined based on the experiment, antibody stability, and optimal storage conditions. Conjugated antibodies should be stored away from light.
