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ReadiLink™ Rapid XFD555 Antibody Labeling Kit *XFD555 Same Structure to Alexa Fluor™ 555*
XFD555 is manufactured by AAT Bioquest, and it has the same chemical structure of Alexa Fluor® 555 (Alexa Fluor® is the trademark of ThermoFisher). Readilink™ protein labeling technology is the most robust and convenient tool for preparing fluorescent antibody conjugates used for fluorescence imaging and flow cytometry applications. ReadiLink™ Rapid XFD555 Antibody Labeling Kit provides the most convenient tool for making XFD555-labeled antibody conjugates. XFD555 dye used in this ReadiLink™ kit is reasonably stable and shows good reactivity and selectivity with protein amino groups. The kit has all the essential components for labeling ~2x50 ug antibody. Each of the two vials of XFD555 dye provided in the kit is optimized for labeling ~50 µg antibody. ReadiLink™ Rapid XFD555 Antibody Labeling Kit requires minimal hands-on time. The prepared XFD555 conjugates (with the kit) are ready to use for fluorescence imaging and flow cytometry applications without further purifications needed. It provides the most convenient method to prepare XFD555-labeled antibodies.
Example protocol
AT A GLANCE
Important
Warm all the components and centrifuge the vials briefly before opening, and immediately prepare the required solutions before starting your conjugation. The following protocol is for recommendation.PREPARATION OF WORKING SOLUTION
Protein working solution (Solution A)
For labeling 50 µg of protein (assuming the target protein concentration is 1 mg/mL), mix 5 µL (10% of the total reaction volume) of Reaction Buffer (Component B) with 50 µL of the target protein solution.Note If you have a different protein concentration, adjust the protein volume accordingly to make ~50 µg of protein available for your labeling reaction.
Note For labeling 100 µg of protein (assuming the target protein concentration is 1 mg/mL), mix 10 µL (10% of the total reaction volume) of Reaction Buffer (Component B) with 100 µL of the target protein solution.
Note The protein should be dissolved in 1X phosphate buffered saline (PBS), pH 7.2 - 7.4; if the protein is dissolved in glycine buffer, it must be dialyzed against 1X PBS, pH 7.2 - 7.4, or use ReadiUse™ Bio-Gel P-6 spin column (cat# 60500 from AAT Bioquest) to remove free amines or ammonium salts (such as ammonium sulfate and ammonium acetate) that are widely used for protein precipitation.
Note Impure antibodies or antibodies stabilized with bovine serum albumin (BSA) or gelatin will not be labeled well.
Note For optimal labeling efficiency, a final protein concentration range of 1 - 2 mg/mL is recommended, with a significantly reduced conjugation efficiency at less than 1 mg/mL.
SAMPLE EXPERIMENTAL PROTOCOL
Run conjugation reaction
- Add the protein working solution (Solution A) to ONE vial of labeling dye (Component A-XFD555), and mix them well by repeatedly pipetting for a few times or vortex the vial for a few seconds.
Note If labeling 100 µg of protein, use both vials (Component A) of labeling dye by dividing the 100 µg of protein into 2 x 50 µg of protein and reacting each 50 µg of protein with one vial of labeling dye. Then combine both vials for the next step. - Keep the conjugation reaction mixture at room temperature for 30 - 60 minutes.
Note The conjugation reaction mixture can be rotated or shaken for longer time if desired.
Stop Conjugation reaction
- Add 5 µL (for 50 µg protein) or 10 µL (for 100 µg protein) which is 10% of the total reaction volume of TQ™-Dyed Quench Buffer (Component C) into the conjugation reaction mixture; mix well.
- Incubate at room temperature for 10 minutes. The labeled protein (antibody) is now ready to use.
Storage of Protein Conjugate
The protein conjugate should be stored at > 0.5 mg/mL in the presence of a carrier protein (e.g., 0.1% bovine serum albumin). For longer storage, the protein conjugates could be lyophilized or divided into single-used aliquots and stored at ≤ –20°C.Spectrum
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References
View all 43 references: Citation Explorer
Cholinergic and adrenergic innervation of the pancreas in chinchilla (Chinchilla Laniger Molina).
Authors: Radzimirska, Malgorzata and Kuchinka, Jacek and Nowak, Elzbieta and Trybus, Wojciech and Szczurkowski, Aleksander
Journal: Folia histochemica et cytobiologica (2020): 54-60
Authors: Radzimirska, Malgorzata and Kuchinka, Jacek and Nowak, Elzbieta and Trybus, Wojciech and Szczurkowski, Aleksander
Journal: Folia histochemica et cytobiologica (2020): 54-60
When the air hits your brain: decreased arterial pulsatility after craniectomy leading to impaired glymphatic flow.
Authors: Plog, Benjamin A and Lou, Nanhong and Pierre, Clifford A and Cove, Alex and Kenney, H Mark and Hitomi, Emi and Kang, Hongyi and Iliff, Jeffrey J and Zeppenfeld, Douglas M and Nedergaard, Maiken and Vates, G Edward
Journal: Journal of neurosurgery (2019): 1-14
Authors: Plog, Benjamin A and Lou, Nanhong and Pierre, Clifford A and Cove, Alex and Kenney, H Mark and Hitomi, Emi and Kang, Hongyi and Iliff, Jeffrey J and Zeppenfeld, Douglas M and Nedergaard, Maiken and Vates, G Edward
Journal: Journal of neurosurgery (2019): 1-14
Efficient Long-Range, Directional Energy Transfer through DNA-Templated Dye Aggregates.
Authors: Zhou, Xu and Mandal, Sarthak and Jiang, Shuoxing and Lin, Su and Yang, Jianzhong and Liu, Yan and Whitten, David G and Woodbury, Neal W and Yan, Hao
Journal: Journal of the American Chemical Society (2019): 8473-8481
Authors: Zhou, Xu and Mandal, Sarthak and Jiang, Shuoxing and Lin, Su and Yang, Jianzhong and Liu, Yan and Whitten, David G and Woodbury, Neal W and Yan, Hao
Journal: Journal of the American Chemical Society (2019): 8473-8481
Intracerebroventricular Delivery of Recombinant NAMPT Deters Inflammation and Protects Against Cerebral Ischemia.
Authors: Chen, Fenghua and Weng, Zhongfang and Xia, Qinghai and Cao, Catherine and Leak, Rehana K and Han, Lihong and Xiao, Jian and Graham, Steven H and Cao, Guodong
Journal: Translational stroke research (2019): 719-728
Authors: Chen, Fenghua and Weng, Zhongfang and Xia, Qinghai and Cao, Catherine and Leak, Rehana K and Han, Lihong and Xiao, Jian and Graham, Steven H and Cao, Guodong
Journal: Translational stroke research (2019): 719-728
Enhanced broadband fluorescence detection of nucleic acids using multipolar gap-plasmons on biomimetic Au metasurfaces.
Authors: Narasimhan, Vinayak and Siddique, Radwanul Hasan and Hoffmann, Magnus and Kumar, Shailabh and Choo, Hyuck
Journal: Nanoscale (2019): 13750-13757
Authors: Narasimhan, Vinayak and Siddique, Radwanul Hasan and Hoffmann, Magnus and Kumar, Shailabh and Choo, Hyuck
Journal: Nanoscale (2019): 13750-13757
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