Product nameAlexa Fluor® 647 Conjugation Kit (Fast) - Lightning-Link®
Alexa Fluor® 647 Conjugation Kit / Alexa Fluor® 647 Labeling Kit ab269823 uses a simple and quick process for Alexa Fluor® 647 labeling / conjugation of antibodies. It can also be used to conjugate other proteins or peptides. Learn about our antibody labeling kits and their advantages.
To conjugate an antibody to Alexa Fluor® 647 using this kit:
- add modifier to antibody and incubate for 15 mins
- add quencher and incubate for 5 mins
The Alexa Fluor® 647 conjugated antibody can be used immediately in WB, ELISA, IHC etc. No further purification is required and 100% of the antibody is recovered for use.
Learn about buffer compatibility below; for incompatible buffers and low antibody concentrations, use our rapid antibody purification and concentration kits. Use the FAQ to learn more about the technology, or about conjugating other proteins and peptides to Alexa Fluor® 647.
Custom size conjugation kits up to 100 mg are available on demand. Please contact us to discuss your requirements.
This product is manufactured by Expedeon, an Abcam company, and was previously called Lightning-Link® Rapid Alexa Fluor® 647 Labeling Kit. 337-0005 is the same as the 100 µg size. 336-0010 is the same as the 3 x 100 ug size. 336-0030 is the same as the 3 x 10 ug size. 336-0005 is the same as the 100 µg size. 336-0015 is the same as the 1 mg size.
Amount and volume of antibody for conjugation to Alexa Fluor® 647
Kit size Recommended
amount of antibody1
amount of antibody
3 x 10 µg 3 x 10 µg 3 x 20 µg 3 x 10 µL 100 µg 100 µg 200 µg 100 µL 3 x 100 µg 3 x 100 µg 3 x 200 µg 3 x 100 µL 1 mg 1 x 1 mg 1 x 2 mg 1 x 1 mL
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.
Buffer Requirements for Conjugation
Buffer should be pH 6.5-8.5.
Compatible buffer constituents
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.
50mM / 0.6% Tris1 0.1% BSA2 50% glycerol 0.1% sodium azide PBS Potassium phosphate Sodium chloride HEPES Sucrose Sodium citrate EDTA Trehalose
1 Tris buffered saline is almost always ≤ 50 mM / 0.6%
2 BSA can also interfere with the use of the conjugated antibody in tissue staining.
Incompatible buffer constituents
Thiomerosal Proclin Glycine Arginine Glutathione DTT
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.
Storing and handling conjugation kits
Lyophilized Lightning-Link® components are hygroscopic.
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.
Storage instructionsStore at -20°C. Please refer to protocols.
Components 100 µg 1 mg 3 x 10 µg 3 x 100 µg ab274049 - Alexa Fluor 647 1 x 100µg 1 x 1mg 3 x 10µg 3 x 100µg ab273994 - Modifier reagent 1 x 200µl 1 x 200µl 1 x 200µl 1 x 200µl ab273995 - Quencher reagent 1 x 200µl 1 x 200µl 1 x 200µl 1 x 200µl
Yuan, Yue, et al used Alexa Fluor 647 Conjugation Kit (Fast) - Lightning-Link (ab269823) as part of examining changes in the nanoscale organisation of CD4 on the surface of CD4+ T cells following HIV-1 binding. They used the kit to conjugate Alexa Fluor 647 to anti-CD4 antibody (OKT4) for use in single-molecule super-resolution imaging.
Representative TIRF-STORM images, and selected magnified regions (insets) of cell-surface CD4 (green) and HIV p24 (magenta). Scale bar = 2 ?m.
Kim, Jin-Wook et al. used Alexa Fluor® 647 Conjugation Kit Lightning-Link® as part of examining alterations in metastatic capacity through cathepsin A by leptin. They used the kit to conjugate anti-LAMP2a antibody for use in immunocytochemistry.
Confocal microscopy images of negative control CHMp cells. (a) DAPI stained CHMp cells; (b) Negative control image of Alexa 488?conjugated secondary antibody. (c,d) Negative control image of Alexa 647 - conjugated LAMP2a antibodies. The white arrow?heads indicate the location of LAMP2a only. The images are captured under 40X confocal microscope.
Thiriet, Pierre-Emmanuel, et al used Alexa Fluor® 647 Conjugation Kit (Fast) - Lightning-Link® (ab269823) as part of rapid and accurate diagnosis of Acute kidney injury (AKI). They used the kit to conjugate Alexa Fluor® 647 to monoclonal anti-Lipocalin-2/NGAL antibody for use in the development of a microfluidic analytical device .
(a-c) Chip description and operation. (a) Presentation of the chip layout. Beads and reagents can be successively injected through the two inlets visible on the left. The device consists of three incubation lines upstream and three concentration lines downstream, at the end of which the beads are accumulated in clusters (shown here in red). (b) Illustration of a sandwich immunoassay used for detection of biomarkers. The analyte we aimed to detect was captured by the bead decorated with capture antibody (cAb) and detection was performed thanks to the fluorescently labeled detection antibody (dAb). (c) Presentation of the successive steps performed on-chip to operate the platform, namely, (1) beads' loading, (2) incubation with detection antibodies and (3) release from the incubation line, (4) clustering in the concentration region, and (5) discarding through the outlet. For the sake of clarity, the species bound to the beads and the electrically activated arrays of electrodes are indicated for each step. (d-f) On-chip incubation of Neutrophil Gelatinase-Associated Lipocalin (NGAL) biomarker. (d) Observation of the small beads' clusters (circled in pink) before and after 15 min of incubation. The fluorescence signal arose from the binding of dAb-NGAL complex to cAb-decorated beads dielectrically trapped in the regions upstream to the electrode line. Three NGAL concentrations were injected in separate experiments, namely, 1 ng/mL, 10 ng/mL, and 100 ng/mL. (f) Fluorescence signal as a function of the incubation time for different NGAL concentrations. After 15 min all concentrations provided a signal greater than the control experiment, consisting of an injection of a solution in absence of NGAL molecules. The error bars were obtained by measuring the fluorescent signal from 10 clusters.
ab269823 has been referenced in 10 publications.
- Yuan Y et al. Single-Molecule Super-Resolution Imaging of T-Cell Plasma Membrane CD4 Redistribution upon HIV-1 Binding. Viruses 13:N/A (2021). PubMed: 33478139
- Lainšcek D et al. A Nanoscaffolded Spike-RBD Vaccine Provides Protection against SARS-CoV-2 with Minimal Anti-Scaffold Response. Vaccines (Basel) 9:N/A (2021). PubMed: 33925446
- Jiao X et al. Generation and Characterization of Typhoid Toxin-Neutralizing Human Monoclonal Antibodies. Infect Immun 88:N/A (2020). PubMed: 32661121
- Laczkó D et al. A Single Immunization with Nucleoside-Modified mRNA Vaccines Elicits Strong Cellular and Humoral Immune Responses against SARS-CoV-2 in Mice. Immunity 53:724-732.e7 (2020). PubMed: 32783919
- Watts LP et al. The RIF1-long splice variant promotes G1 phase 53BP1 nuclear bodies to protect against replication stress. Elife 9:N/A (2020). PubMed: 33141022
- Kim JW et al. Leptin Modulates the Metastasis of Canine Inflammatory Mammary Adenocarcinoma Cells Through Downregulation of Lysosomal Protective Protein Cathepsin A (CTSA). Int J Mol Sci 21:N/A (2020). PubMed: 33255835
- Lederer K et al. SARS-CoV-2 mRNA Vaccines Foster Potent Antigen-Specific Germinal Center Responses Associated with Neutralizing Antibody Generation. Immunity 53:1281-1295.e5 (2020). PubMed: 33296685
- Thiriet PE et al. Rapid Multianalyte Microfluidic Homogeneous Immunoassay on Electrokinetically Driven Beads. Biosensors (Basel) 10:N/A (2020). PubMed: 33371213
- Galeotti C et al. Intravenous immunoglobulin induces IL-4 in human basophils by signaling through surface-bound IgE. J Allergy Clin Immunol 144:524-535.e8 (2019). PubMed: 30529242
- Boonpiyathad T et al. High-dose bee venom exposure induces similar tolerogenic B-cell responses in allergic patients and healthy beekeepers. Allergy 72:407-415 (2017). PubMed: 27341567